MTC Module 1





  • study of disease
  • clinical manifistations(signs, sympt, lab findings)
  • structural changes
  • molecular chanages
  • how do molecular changes result in observed clinical manisfestations




Hutchinson-Gilford Progeria Syndrome(HGPS)


  • baldness
  • swollen veins
  • absence of eyebrowns and eyelashes
  • beak shaped nose
  • shrunken chin
  • narrow chest and swollen and stiff joints
  • narrowing of coronary arteries(premature cardiac disease)



Hutchington-Gilford Progeria Syndrome (HGPS)



  • abnormal lobulated nuclei
  • lack of usual heterochromatin clumping on edge
  • thick electron dense area adjacent to the nuclear membrane where lamins are present
  • abnormal amounts of lamin A (LMNA gene) are present





  • an intermediate filament that polymerizes along the inner nuclear membrane which has a structural and regulatory role





  • an farnesyltransferase inhibitor that blocks production of farnysl-progerin
  • developed as a cancer chemotherapy agent but has some effect in-vitro and in phase 1 and 2 of clinical trials
  • used to treat HGPS




Treatment of HGPS

  • farnesyltransferase inhibitor (FTI) like inofarnib
  • using small, interfering RNA molecules to interfere with the transcription of the mutant allele
  • siRNA exon skipping might also be useful to eliminate the site to which the farnesyl moiety is bound




Amino Acid Structure

  • central ?-carbon
  • carboxyl group (coo-)
  • amino group (NH3+)
  • hydrogen
  • Side chain (R)




Chirality of Amino Acids

  • L-amino acids the hydrogen is on the right
  • D amino acid the Hydrogen is on the left
  • all chiral except glycine
  • most mamalian amino acids we are interested in are L amino acids




Peptide Bond Formation

  • condensation reaction btw COO- and NH3+
  • usually R groups are in trans- location where one is up and the others down
  • if they can interact and structure allows it then they can be in cis?




Weak Acid Vs Strong Acid

  • weak acid is incomplete dissociation in water
  • strong acid is complete dissociation in water




Henderson-Hasselbalch Equation

  • pH=pKa+log10 [A-]/[HA]




pKa of ?COOH and ?NH3

  • ?COOH = 1.8
  • ?NH3 = 9.3




Amino Acid R-Group


  • hydrophobic interaction (ex phenylalanine side chains)
  • hydrogen bonds (H-O usually)
  • electrostatic interactions or ionic bonds(salt bridges)




Primary Structure

  • order of amino acid residues in a polypeptide chain or protein
  • determined by the sequence of codons in mRNA resulting from genomic transcription and splicing




Start Codon

  • AUG
  • methionine 
  • usually cleaved off by enzymes after translation




Non-critical variation of structure

  • these are called hidden variations
  • this is often the case with conservative substitution(amino acids that are similar
  • same function and same structure




Critical to structure variation

  • different structure and different function
  • increases activity or decreases activity
  • or alters structural integrity (may  be subject to cellular proteolysis)





  • sequence variations in an allele within a population
  • sometimes important sometimes not 




Developmental Variation

  • certain variants of a protein family are differentially expressed based on the developmental stage of the organism
  • the variant proteins are called isoforms or isozymes
  • example are different forms of hemoglobin that have different affinities for O2



Post-translation Mod.

Carbohydrate addition

  • o-glycosylation: OH of ser, thr, tyr
  • N-glycosylation: NH2 of asn




Post-translation Mod.

Lipid Addition

  • palmitoylation: internal SH of cys
  • myristoylation: NH of N-terminal gly
  • Prenylation: SH of cys



Post-translation Mod.


  • phosphorylation: OH of ser, thr, tyr
  • acetylation: NH2 of lys, terminus
  • ADP-ribosylation: N of arg, gln; S of cys




Post-translation Mod.

Modified amino acids

  • oxidation: pro, lys
  • carboxylation: glu




Secondary structure

  • alpha helix (right handed) all r groups on the outside?
  • beta pleated sheet is either antiparallel or parallel (can stack on one another)
  • beta turn




Tertiary Structure

  • patterns of secondary structure
  • structural motif->subdomain->domain->fold
  • 1393 folds identifield 
  • no new folds discovered since 2008
  • nucleotide binding fold, actin fold ect 




Quaternary Structure

  • the specific interaction of distinct polypeptide chain subunits (geometrically stoichiometrically)
  • means to produce fully functional highly stable proteins




Structural forms of proteins

  • globular
  • fibrous
  • membrane spanning




Factors influencing protein folding

  • mutations
  • environment(synthesis, functional site-ph, salt, temp, oxidizers)
  • chemical modification (enzymatic and non-enzymatic)
  • protein-protein interactions (chaperones, other interacting proteins, abnormal proteins)





  •  heme porphyrin 9 group found in hemoglobin (4 tetraporphyrin groups)
  • iron in center (Fe2+)
  • iron bound by 5 Nitrogens and one oxygen
  • oxygen pulls heme group into plane with the iron and moves the helix that the heme is attached too (cooperativity) 
  • attached to hemoglobin or myoglobin by proximal histidine





  • a lot of alpha helical structure 
  • single monomer with one heme group binded by proximal histidine 
  • not great at grabbing oxygen nor releasing it
  • found in muscles in high amounts





  • has two alpha and two beta subunits (quaternary structure) 4 protomers(monomers)
  • each protomer has its own heme group
  • participates in cooperativity(easier to bind with every new oxygen bound)
  • means multiple affinities for oxygen(sigmoidal curve)
  • T(tense) state and R(resting) state
  • if no cooperativity the curve would shift to way right and wouldnt get to 1.0 and not be sigmoid





  • allows for hemoglobin cooperativity
  • called bisphosphoglycerate because PO4 groups on different carbons(interacts with His and Lys
  • inc BPG a right shift of sigmoid curve happens (takes higher pO2 to get 1/2 O2 sat.)
  • dec BPG a left shift happens to sigmoid curve(takes lower pO2 to get 1/2 O2 sat.)




Fetal Hemoglobin

  • O2 flows from maternal hemoglobin to fetal hemoglobin
  • fetal hemoglobin does not bind BPG  so it has a leftward shift, so can pick up oxygen more readily from lower O2 environment




pH and Hemoglobin Binding Curves

  • lower the pH (more H+), the more to the right the curve is so it releases its oxygen at high pO2 levels
  • the higher the pH (less H+) the more to the left the curve so it releases its oxygen at lower pO2 levels




Bohr Effect

  • a decrease in blood pH and increase in CO2 leads to hemoglobin molecules to release there oxygen more readily with the oposite being true
  • this is causes because an increase in CO2 reacts with water to form carbonic acid, increasing pH
  • an added proton(H+) from low pH  stabilizes the ionic bonds favoring the Tstate of Hb and release oxygen
  • the high pH removes the H+ from the Hb(in lungs)




CO2 transport to Lungs

  • bulk of it is transported by bicarbonate H2CO3
  • some of it is transported by carbamate of hemoglobin
  • Hb-N-COO- + H+
  •        |
  •        H




Hb Curve Right Shift

  • caused by increase in pCO2, temperature, acidity(lower pH) or BPG




Carbon Monoxide

  • binds to iron in heme at a much greater affinity then oxygen
  • takes long time to come off 
  • if one CO is bound the Hb holds onto its oxygens 




Detection of Abnormal Hemoglobins

  • hb genes are expressed co-dominant hence normal and abnormal may be present
  • a change in AA sequence can result in a change in charge so physical seperation can be used (electrophoresis or chromatography)
  • can be performed on meltimers or on seperated chains
  • DNA sequencing can then be performed
  • common hemoglobinopathies can be quickly detected through simpler techniques like sickle cell preps and blood smears




Hemoglobin S and C


  • hemoglobin S trait (?A?S) is present in 8% of african americans, and as high as 30% of individuals in some areas of africa and middle east
  • hemoglobin C trait (?A?C) is present in 5% of african americans




6th Amino Acid for 

Hemoglobin S and C

  • A position 6 Glu, isoelectric point is 7.0
  • S position 6 Val, isoelectric point is 7.2
  • C position 6 Lys, isoelectric point is 7.4




Hb and Alkaline Electrophoresis

  • carried out at pH of 8.6 because all Hb are negative at this charge and migrate towards anode
  • the magnitude of the negative charge will determine how rapidly the hemoglobin migrates
  • many hemoglobin variants migrate to the same point




Hb and Acid Electrophoresis

  • performed in agar with a citrate buffer at pH 6.2 and is a complement to alkaline electrophoresis
  • agaropectin in the agar binds reversibly to certain AA on the surface or heme pocket
  • the Hb-agaropectin complex moves based on amount of binding
  • confirsm S and separates C from other Hb




Hb and Isoelectric focusing

  • many ampholytes(small peptides which carry charge and affect pH) are suspended in agarose
  • when current is applied these ampholytes establish a pH gradient across the strip
  • applied hemoglobins migrate to their isoelectric point
  • this yields sharp bands but more minor bands
  • static vs dynamic and widely used in pop. screening




Hb and High Performance Liquid


  • uses a weak cation column, as ionic strength of eluting buffer increases Hb elute with a particular retention time
  • Hb with AA substitutions present will have a different retention time that A(2.5 mins)- for example S(4.48 mins) and C (5.15 mins)




Hemoglobin S

  • deoxygenated hb polymerize and sign polymerization leads to formation of helical filaments
  • Hb S has a greater tendency to polymerize
  • RBC with S filaments are rigid (sickle) (not elastic)
  • removed from circ causing an anemia and RBC breakdown products
  • occlude small vessels causing ischemic injury




Hemoglobin C

  • less soluble then Hb A
  • tends to crystalize in the cyell
  • crystal formation is more likely in the oxygenated state
  • formation of crystals decreases the life span of the RBC
  • patients with only C Hb have an increased red cell turnover but usually only mild anemia





  • shortage of blood supply to tissues, causing a shortage of oxygen and glucose needed for cellular metabolism





  • results from an altered expression of one of the globin chains
  • when there is a significant lack of expression of a globin chain, alternate normal Hb or alternate Hb multimers are present
  • in beta-thalassemia-Hb A is decreased and Hb A2(delta) and F(gamma) chains are expressed in increased amounts
  • in alpha-thalassemia- all Hb contain alpha chains so abnormal multimers are present-Hb H and Hb Barts are seen (both real unstable)




Structural Hb changes that effect

loading and unloading

  • FC corner(?94-104) changes affect change configuration movement association with uptake-unloading
  • C terminus (? 143-146) change affect DPG binding site





  • increase in hemoglobin levels
  • can be due to pathologic or physiologic state
  • pathologic-erythropoietin secreting tumor(or adm of erythropoietin)
  • physiologic hypoxia-living at high altitude
  • pathologic hypoxia-chronic lung disease
  • pathologic failure to release oxygen from hemoglobin




Hb Andrew-Minneapolis

  • lys>Arg substitution in AA 144 in ? globin chain
  • P50 is 18mm Hg, Loss of 2,3 DPG effect
  • variant hemoglobin is always heterozygote state(homozygous state is assumed to be lethal




Hb Malmo

  • FG corner Variant-His>Gln AA change in ? chain at position 97
  • described in southern sweden
  • does not show up on electrophoresis
  • increased oxygen binding affinity (does not release at high pO2)




1st order kinetics

  • at substrate concentrations below Km (affinity for enzyme)
  • reaction rate proportional to substrate
  • many drugs are metabolized in liver where drug concentrations are below Km but sometimes excessive alcohol ingestion can saturate the alcohol dehydrogenase  




Zero-Order Kinetics 

  • high substrate concentration reaction
  • reaction is independent of substrate
  • all enzyme is in [ES] and not limited by [S], but by the turnover number of the enzyme [Kcat]




2nd Order Rate Reaction

  • dependent on two substrates, substrate and cofactor
  • thus doubling the concentration of one substrate doubles the rate
  • while doubling both increases the rate 4-fold
  • cofactor is either vitamin or coenzyme




2nd Order Reaction Enzyme

  • active site contains unique amino acid domain and cofactor that form covalent bonds with substrate
  • formation of the transition state requires energy for covalent bond formation
  • energy derived from ionic, h bonds, hydrophobic, fold free energy of enzyme in ES
  • once energy barrier is overcome, rx proceeds w/o additional energy




Substrate binding site of enzyme

  • amino acids of the substrate binding site determine what substrate binds
  • 2 models: lock and key model and induced fit model
  • both models suggest that, upon ubstrate bidning, the enzyme active site changes conformation the the transition state




Lock and Key Model

  • model for substrate binding site of enzyme
  • amino acid domain of active site determimes the type of substrate bound through hydrophobic, electrostatic(ionic) or hydrogen bonds
  • ex is glucokinase that binds glucose with ionic bonds and hydrogen bonds




Induced fit model

  • model for substrate binding site of enzyme
  • substrate binding induces conformational change necessary for hydrophobic, ionic, and hydrophillic bond formation




Transition State Complex

  • substrate activation requires energy for the formation of covalent bond formation of the transition state complex
  • enzymes increase the rate of reaction,kcat, by decreasing the activation energy through either acid-base or electronic stabilization of the [ES]




Transition State Analog Inhibitors

  • highly potent and selective enzyme inhibitors because they mimic the three dimensional structure of the substrate transition state with pre-formed covalent bonds and thus require no energy of activation
  • bind enzymes a million fold tighter then substrate, and thus inhibitors can be administered at lower concentrations to avoid adverse drug toxicity
  • have high affinity(low Km)




Serine Protease

  • cleaves preproteins(zymogens) to make them active
  • pivotal role in blood coagulation(prothrombin) and digestion(trypsin, chymotrypsin and elastase)
  • hydrolyze peptide bonds on the carboxyl side of hydrophobic amino acids, Phe, tyr, trp and leu
  • dissociation of water to hydroxyl group(nucleophile) can attack the protein carbonyl group forming an oxyanion transition state complex
  • hydroxl addition to carbonyl froms a carboxylic groups, and the proton forms an amine




Chymotrypsin Catalysis

  • catalytic triad(Asp-His-Ser) Asp transfers e- to His nitrogen to initiate acid-base catalysis
  • removal of serine H+ makes serine oxygen a highly reactive nucleophile, making oxyanion tetrahedral transition state
  • transition state stabilized by covalent bonds with ser and his functional groups and substrate
  • acyl enzyme interm. serine covalent catalysis
  • water donates e- to carbonyl forming 2nd transition state
  • electron rich tetrahedral draws His H+ to serine
  • oxyanion base draws the his hydrogen(acid), releases acyl-enzyme and reforms OH serine




Thiamine Pyrophosphate (TPP)

  • activation-transfer coenzyme by forming a covalent bond with substrate(pyravate)
  • enzyme function amino acid groups rmove a proton from TPP forming a highly reactive carbanion that attacks positively charged carbonyl group carbon causing bond scission




Coenzyme A

  • synthesized from vitamin pantothenic acid functions to add CoA to fatty acids and cholesterol trapping the in cells
  • in this case instead of bond scission, acyl-CoA synthetase forms a ond btw CoA and fatty acid (acyl) carboxyl group




Vitamin Biotin

  • functions in carboxylase reactions by attaching CO2 to substrate, such as, pyruvate carboxylase converting pyruvate to oxaloacetate
  • forms covalent bond btw itself and substrate
  • forms an activation transfer action




Pyridoxal Phosphate (Viatamin B6)

  • reactive aldehyde group binds amino acids and thus is critical for aminotransferase enzyme activity




Activation Transfer cofactor features

  • coenzyme functional group binds enzyme 
  • coenzyme reactive group forms or breaks covalent bond
  • coenzyme can be thought of as functioning in 2nd order kinetic reactions since they bind enzyme substrate binding site




Coenzymes in Oxidation Reduction 


  • co-enzymes NAD+, FAD+, FMN and NADP+ function in the transfer of electron and hydrogen from substrate by donating or accepting eletron as hydride, hydrogen or oxygen
  • unlike act. transfer coenzymes, oxi-red coenzymes do not form covalent bonds with substrate
  • lactate dhase binds NAD+ thru AMP moiety and NAD+ functional group accepts hydride ion oxidizing lactate to pyruvate




NAD+ and FAD

  • coenzymes in oxidation reduction catalysis
  • NAD+ is synthesized from the vitamin Niacin and FMN
  • FAD is synthesized from Riboflavin
  • function in the transfer of electron and hydrogen from substrate by donating or accepting electrons




Vitamins C and E

  • function in oxidation-reduction reactions and as antioxidants by donating electrons(reducing agents)




Metal Ions

(Enzyme Catalysis)

  • metal ions have a positive charge and thus function as electrophile(electron loving)
  • assist in substrate binding to enzyme, stabilization of function groups in acid-base catalysis, accepting and donating electrons in oxidation-reduction reactions
  • alcohol dhase(ADH) catalyzed oxidation of alcohol to acetaldehyde requires zinc to stabilize serine hydroxyl group that removes a proton from alcohol leaving serine oxyanion stabilized by Zn2+, and thus allowing hydride transfer to NAD+




Covalent Enzyme Inhibitors

  • form tight bonds with function group amino acids of the catalytic site
  • usually toxins or selective drugs
  • usually irreversible inhibitors and thus require new enzyme synthesis
  • examples are aspirin and 5-fluorouracil





  • covalent coenzyme inhibitor
  • irreversible inhibitor of prostaglandin endoperoxide synthase (PGH)
  • PGH has endoperoxidase and cycloxygenase activity (COX) and normally abreviated as COX1 and COX2





  • chemotherapeutic drug
  • covalent enzyme inhibitor
  • inhibits thymidylate synthase in the conversion of dUMP to dTMP for DNA synthesis




Competitive Inhibitors

  • do not change Vmax because inhibitor binding is reversible and can be overcome by high substrate concentrations
  • however Km is increased becasue sub binding impairment
  • structurally related to normal substrate
  • product inhibition by products is an important mech of feedback inhibiton: methanol and ethanol competes for alcohol dhase and methanol goes to formaldehyde?
  • succinate dhase metabolizes succinate to fumarate, which is comp. inhibited by malonate




Non-Competitive Enzyme Inhibitors


  • structurally unrelated to substrate
  • bind to enzyme, not at catalytic site, to inhibit ES and E
  • decrease Vmax with out changing Km
  • cyanide, heav metal ions and some insecticides are all nc inhibitors
  • Na/k ATPase is inhibited by ouabain
  • lead forms covalent bonds with sulfhydryl side chain of cystein, causing nc inhibition in things like ferrochelatse 




Un-competitive Inhibitors 


  • bind only to the enzyme substrate complex and not the free enzyme
  • decrease both Vmax and Km
  • unlike comp inhibitors, uncompetitive inhibitors work best at high substrate concentrations




Suicidal or Irreversible inhibitors

  • irreversible inhibitors bind covalently and only way to overcome inhibition is create new enzyme
  • also called mixed inhibitors because they bind both E and ES and decrease Vmax and do not change Km
  • most irreversible inhibitors are environmental toxins or drugs MAO inhibitors allow neurotransmitter to remain active longer
  • organophasphate (acetylcholinesterase), allopurinol (gout/xanthine oxidase) and aspirin(cyclooxygenase)




Transition State Analogs

  • resemble enzyme transition state substrate
  • penicillin binds glycopeptidyltransferase, a bacteria enzyme required for cell wall synthesis
  • partial reaction of penicillin transition state analog with the enzyme results in covalent bond formation and suicide inactivate of enzyme
  • completely inactive enzyme activity irreversibly




Transition State Analog Inhibitors(TSAI)

  • have been synthesized and are presently used in clinical trials
  • mimic substrate enzyme transition state and thus do not require energy to form covalent bonds, thus they bind with high specificity and 1 million fold more tightly to enzyme than substrate
  • high affinity and avidity reduce druge dose reducing adverse effects





  • TSAI requires covalent bond to inhibit xanthine oxidase and uric acid synthesis from purine nucleosides
  • allopurinol is converted by xanthine oxidase to its transition state analog oxypurinol that irreversibly inhibits enzyme activity, suicide inactivation





  • inhibits conversion GMP to guanine to
  • instead converts to GTP and then to dGTP that causes T-cell Apoptosis
  • inhibits Purine nucleotide phosphorylase(PNP) that functiosn to remove ribose 1-phosphate from purine nucleoside producing the base that can be metabolized to uric acid





  • most common enzymes involved in metabolism that transfers electrons from substrate to co-enzyme NAD+, FAD+, NADP+, or elecrons from coenzyme to the substrate
  • the enzymes, dehydrogenase, are usually named for the substrate they metabolize





  • oxido-reductase enzymes
  • use molecular oxygen as a substrate and normally a co-enzyme for activity





  • oxido-reductase enzymes
  • cytochrome P450 
  • transfer one molecule of oxygen to substrate and other to water
  • found in reduction of steriods and eliminate of toxic drugs and conversion of drugs





  • oxido-reductase enzymes
  • such as COX enzyme 
  • insert two molecules of oxygen into substrate





  • oxido-reductase enzymes
  • transfer electrons from the substrate to oxygen producing hydrogen peroxide
  • peroxidase convert hydrogen peroxide to water, therefore oxidase can have two activities





  • catalyze the group transfer reactions
  • if the transfer group is a high energy group, the enzyme is a kinase
  • glycosyltransferase transfers carbohydrate while acyltransferase transfers fatty acids
  • transaminations are important for measuring blood to determine organ damage
  • synthase also function in transfer of glucose from UDP or ehtanolamine from CDP and unlike synethase it does not require ATP





  • cleaves C-O, C-N, or C-S bonds by addition of water
  • most digestion and lysosomal ezymes are hydrolases (chymotrypsin)
  • hydrolase names indicate the substrate or bonds on which they act, which include petidase, lipases, esterases, and phosphatases





  • remove a group nonhydrolytically forming a double bond such as with dhydratase 
  • lyases cleave C-C, C-O, and C-N bond by means other than hydrolysis or oxidation
  • are many times reversible, creating a new bond and obliterating a double bond 
  • when they favor the formation of a single C-C bond they are often called synthase





  • interconvtert positional, geometric or optical isomers
  • isomerases that catalyze the movement of a phosphate from one atom to another are called mutases
  • isomers are molecules with same formula but different structures




Ligase Enzyme Catalysis

  • in contrast to hydrolase, ligase synthesize C-C, C-S, C-O, C-S, bonds by cleavage of a high energy bond
  • carboxylase addes CO2 to substrate with energy provided by ATP cleavage
  • Synthetase such as fatty acyl-CoA synthetase attaches CoA to fatty Acid by cleavage of a high energy bond




Lineweaver-Burk and Michaelis Menten

  • michaelis-menten is the graph that shows the Km and Vmax and is all in the second quadrant
  • lineweaver-burk takes the equation for that and makes it linear, making 1/vmax the y intercept and -1/Km the x intercept




Hill Plot

  • used when a velocity vs substraight curve is sigmoidal in enzyme kinetics
  • it approximates what a michaelis-menton curve would look like on it, figure out wheres 1/2Vmax is and then keep going over to that point on the sigmoid curve
  • this point is the S0.5 or K0.5




Monomeric G-Proteins


  • when GTP is bound the conformation of Gprotein allows it to bind target proteins, which are then activated
  • gprotein hydrolyzes a phosphate from GTP to form GDP, which changes gprotein confromation and causes it to dissociate from the target protein
  • GDP is exchanged for GTP, which reactivates the gprotein




Glycogen Phosphorylase

and AMP

  • AMP binds to the allosteric site on glycogen b, making it active and ready to accept two phophate groups, one per catalytic subunit 





  • internal cellular proteases involved in driving apoptosis(programmed cell death
  • proteolytic cleavage example





  • class of inactive precursor proteaseswhich prevent them from being active until needed
  • ex. are firbinogen to fibrin and prothrombin to thrombin
  • such pre-enzymes are held in reserve until needed and can quickly become enzymes withough gene expression and protein synthesis




Regulation of Metabolic Pathways

  • regulation occurs at the rate-limiting step
  • feedback regulation
  • feed forward regulation
  • tisse isozmes of regulatory proteins
  • counter regulation of opposing pathways
  • substrate channeling through compartmentation




Assymetry of Plasma Membrane

  • higher concentration of phosphatidylcholine and sphingomyelin in outer leaflet
  • higher content of phosphatidylethanolamine and phosphatidylserine in inner leaflet
  • phosphatidylinositol(cell signaling) is exclusely located in iner leaflet
  • cis double bonds kink, trans do not(trans fat bad)
  • flippases transfer lipids between leaflets




Lipid Anchored Proteins

  • third class of membrane proteins
  • include proteins bound tot he external membrane surface trhough the glycophosphatidylinositolglycan(GPI) anchor, a lipid covalently attached iself to the membrane
  • uses inositol (which is usually only on the inner leaflet but some cases it is not?)





  • single membrane bound that can replicate by by division (no DNA)
  • oxidative rxs: catalase can degrade H2O2 produced from the first step of beta-oxidation(mit.)
  • oxidation of very long chain fatty acids(>c20)
  • conversion of cholesterol to bile acids
  • originated from ER




Peroxisome Defect


  • Zellweger syndrome, neonatal adrenoleukodystrophy, Infantile Refsum Disease
  • IRD is less severe then the others
  • Xlinked ALD is defect in ABCD1, the VLCFA transporter to peroxisome(accumulation of VLCFA)(leads to death)
  • Refsum disease defect in the alpha oxidation enzme, phytanoyl-coa hdase, or PEX7 leading to phytanic acid accumulation (dieting from dairy and ruminant meat can help)





  • single membrane with internal pH of 5.5
  • membrane proteins protected from digestion y glycosylation
  • responsible for digestion, elimination of things
  • orginated from golgi and endosome
  • lysosomal storage disease is caused by build up of undigested material in lysosome from defect in enzymes (tay-sachs, niemann-pick, gauchers)




Mitochondria DNA Genome

  • encodes 37 genes
  • 13 for subunits of respiratory complexes
  • 22 for mitochondrial tRNA, 2 for rRNA
  • genetic defects lead to complex disease such as mitochondrial encephalomyopathy
  • most mitochondria are maternal
  • MELAS? (possibly a disease)




Smooth Endoplasmic Reticulum


  • Microsomes formed by mechanical fragmentation
  • responsible for lipid synthesis, metabolize drugs and xenobiotich chemicals(cytochromP450) for detoxification, steroid hormone syntehsis
  • glycogen storage in liver cells




Gated Channels

  • transmembrane proteins form a pore for ions that is either opened or closed in response to stimulus
  • may be voltage gated, ligand gated, or a regulator change in the intracellular domain(phosphorylation-gated and pressure gated)
  • Cystic Fibrosis transmembrane conductance regulator(CFTR) is an example of a ligand gated channel controlled by phosphorylation




Caveolae mediated transport

  • potocytosis
  • cave like structure in membrane
  • full of sphinoglipids
  • formed by caveolaen(cholesterol binding protein)
  • lipid raft?




ATP-Binding Cassette

Transporter Superfamily

  • bind ATP and use the energy to transport hydrophobic molecules across cell membranes
  • classified based on the homology in the ATP binding domains
  • consists of 7 mammalian subfamilies based on similarity in gene structure 
  • chart in slides*




Fatty Acid Transporters


  • first fatty acids are freed from dietary fat by gastric lipases and transported across the apical membrane of the intestine actively by FATP4, FAT/CD36, FABPpm, or passively diffuse through lipid bilayer in enterocytes, FABPc facilitates fatty acid transport through the cytosol.
  • majority is re-esterfied to TAG and excreted into circulation as chylomicrons(CM)
  • membrane and non-membrane associated transporters




Fatty Acid Membrane

Associated Transporters

  • plasma membrane fatty acid binding protein coopers with CD36 to transport fatty acids
  • fatty acid translocase/CD36 two transmembrane spanning regions major form responsible forinsuline inducible FA uptake in muscle involved in atherosclerosis
  • fatty acid transport proteins-six trans transmembrane domains 
  • FATP1(white adipose and muscle), FATP2 and 5(hepatocytes) FATP3(many tissues, FATP4 (enterocytes), FATP6(cardiomyocytes




Fatty Acid Non Membrane

Associated TRansporters

  • cytosolic faty acid binding protein(FABPc)
  • facilitate intracellular FA trafficking to targe organelles such as mitochondria or nucleus
  • serum albumin transports FA to target tissues in blood circulation




Glucose Transporters

  • Na+ dependent cotransporter (Na/K pump helps keep sodium gradient to facilitate this)
  • facilitative glucose transporter
  • insulin can stimulate these





  • glucose transporter
  • in human erythrocyte and blood brain barrier
  • expressed in cell types with barrier functions
  • although GLUT1 in erythrocytes has a overall km of 1-7mM, it is abundant to transport serum glucose adequately evin in the hypoglycemic level (2.2mM)





  • glucose transporter
  • found in liver, kidney and pancreatic ?-cell
  • high capacity, low-affinity transporter
  • high Km value of liver GLUT2 allow serum glucose available for other tissues, since liver has its own capacity for glucose production





  • glucose transporter
  • found in brain(neurons)
  • major transporter in the central nervous system
  • exhibits very low Km value for glucose because the brain uses glucose as the only carbohydrate energy source




GLUT 4 and 5

  • glucose transporter
  • GLUT4 found in adipose tissue and skeletal muscle and is insulin sensitive
  • GLUT5 is found in intestinal epithellium and spermatozoa and is a fructose transporter




Types of Chemical Messengers

  • Neurotransmitters(nervous system)
  • hormones(endocrine system)
  • cytokines(immune system)
  • retinoids, eicosanoids, growth factors, ect





  • a neurotransmitter secreted from neurons in response to an electrical stimulus
  • relays a signal from a motor nerve to a muscle fiber
  • when action potential is reached, voltage gated Ca2+ channels open triggering fusion of Ach synaptic vesicles with plasma membrane
  • the release Ach binds to Ach receptors causing its conformational change allowing Na+ to diffuse in and K+ to diffuse out triggering a cellular response
  • signal is terminated by acetylcholinesterase





  • small peptie(4-36 AA) secreted by neurons
  • neuropeptide alpha-MSH is melanocyte stimulating hormone:anorexigenic peptide (controlling appetite)
  • neuropeptide Y:released from hypothalmus (orexigenic peptide)





  • use gprotein coupled receptors
  • control cellular function in response to injury
  • are all derived from arachidonic acid
  • prostaglandins, thromboxanes and leukotrienes




Chemical Messangers for 

Intracellular Transcription factors

  • either hydrophobic or lipophilic
  • mostly gene-specific transcription factors
  • transported in blood bound to serum albumin or toa  more specific transport protein such as steroid homrone binding globulin(SHBG) or thyroid hormone binding globulin (TBG)




Cytoplasmic Steroid/thyroid 

Hormone Receptors

  • asociated with Heat Shock Proteins (HSPs) in absence of ligands (HSP90)
  • binding of ligands triggers HSP release and migration of the receptor into the nuclease to a specific DNA site called the hormone response element
  • ex: PR(progesterone), GR(cortisol), MR(aldosterone)




Nuclear Steroid/Thyroid

Hormone Receptors


  • mostly bound to DNA as a heterodimer with RXR in an inactive state without ligands (most like this)
  • some homodimer or monomer
  • when active ligand bind, the receptor recruit protein complexes which initiate gene transcription
  • ex: PPARs(fatty acids), LXR(oxysterol), FXR(bile salts), VDR (vitamin D2)




Major Classes of Membrane Receptors

  • Ion-Channel Receptors
  • Receptors that are kinases or that bind kinases
  • Heptahelical receptors (Gcoupled Protein Receptors)(7membrane spanning alpha helices)




Tyrosine Kinase Receptors

  • Ras and the MAP kinase Pathway
  • phosphatidylinositol phosphates in signal transduction
  • JAK-STAT Receptors
  • Receptor Serine/Threonine Kinases




Ras and the MAP Kinase Pathway

  •  a growth factor as ligand for the two proteins of this
  • receptor dimerizes which activates tyrosine kinase activity
  • this activity phosphoralates each others receptors
  • recruits Grb2 contains SH2 domain(recognizes phosphorylated tyrosine residues)
  • Also recruits SOS contains proline rich domain that binds to SH3 of Grb2 and GEF that associates with RAS
  • when SOS binds with RAS, RAS exchanges GDP for GTP and becomes activated and goes and activates enzyme Raf (with MAPK)




Phosphatidylinositol Phosphate 

in signal transduction

  • signaling molecules can be generated thru tyrosine kinase receptors or GPCR
  • 2 ways to initiate signaling 
  • 1:phosphatidylinositol 4, 5 bisphosphate is cleaved to generate 2 important intracellular messengers, diacylglycerol and inositol triphosphate (gprotien uses enzyme PLC)
  • both calcium from IP3 and DAG activates PKC
  • 2: PI-4,5-bisP is converted by PI 3-kinase to PI-3,4,5-trisP, which serves as a plasma membrane docking site signal transduction protein containing pleckstrin homology domains




PLC subtypes

  • PLC gamma is activated by tyrosine kinase
  • PLC beta is activated by GPRC




Phosphatase and Tensin Homolog


  • terminates or inhibits PI3K activation via dephosphorylation of PIP3 to PIP2




PI3 Kinase

  • PI3Ks are grouped into 3 classes depending on their structural homology and substrate preference
  • among 3 classes, class 1 subfam. are involved in PIP3 production and form heterodimers consisting of one catalytic subunit and one regulatory subunit
  • their regulatory subunits determine the specificity of membrane-receptor association
  • Class 1A: P85 w/ Rec Tyro Kinase, Class 1B: p101 with GPRC)




Insulin Receptor

  • dimer with each half containing an ? and ? subunit
  • very divergent signaling pathway
  • can activate PLC kinase, PLC gamma, actiavte G protein 




Insulin Receptor

PLC kinase pathway

  • insulin binds and tyrosine activity phosphorylates each half of the dimers domain
  • this phosphorylation recruits Insulin Receptor Substrate (IRS)(has an SH2 Domain)
  • IRS phosphorylated and recruits PI3 kinase(SH2 domain)
  • PI3 kinase converts PIP2 to PIP3
  • PIP3 thend does stuff with PDK 1 and PKB which activates and leads to mTORC1 from mTORC2




JAK-STAT Receptors

  • leptin can bind to these
  • no intrinsic kinase activty
  • binds the tyrosine kinase JAK(janus kinase)
  • these phosphorylate each other
  • they then phosphorylate the JAK-STAT receptor
  • this recruits STATs that are then phosphorylated
  • STATs then dissociate and translocate to the nucleus





  • regulates neuronal activity of satiety in the hypothalamus to control energy balance and body weight
  • induces STAT3 homodimer formation
  • increase in lepten decreases apeteite 




Receptor Serine-Threonine Kinase

  • transforming growth factor ? and bone morphogenetic proteins (BMP)
  • receptor exists as monomer (Type II) but recruits type I receptor after ligand bind
  • type II phosphorylate serine receptor of type I
  • this then phosphoryalates serine residues of receptor(R) Smad
  • this then complex with commom(Co)-Smad and both then translocates to the nucleus





  • protein found in the receptor serine-threonine kinase pathway
  • Receptor-Specific Smads: Smad 1,2,3,5, and 9
  • Common Smad: Smad 4
  • Inhibitory Smads: Smad 6 and 7




Phosphatidylinositol Signaling

  • activation of phospholipase C by G?q hydrolyzes PIP2 into DAG and IP3
  • IP3 stimulates Ca2+ release from the SR
  • DAG activates Protein Kinase C (PKC)




Guanylyl Cyclase Receptors

  • convert GTP to cGMP
  • elevated cGMP activate PKG that phosphorylates target proteins
  • Nitric Oxide Receptor (syn from L arginine)and Atrial natriuretic peptide (ANP) receptor




Atrial Natriuretic Peptide (ANP) Receptor

  • effect guanyly cyclase 
  • membrane bound receptor protein
  • ANP is hormone secreted by heart muscle to decrease blood pressure and 
  • acts on two sites, kidney and vascular smooth muscle
  • triggers renal excretion of Na+ and consequently of water to reduce blood volume and relax blood vessels




cGMP Elevating Drugs

  • effect guanylyl cyclase
  • Glycerol trinitrate: produce NO: for Angina Pectoris
  • Nesiritide(synthetic ANP): For heart failure
  • Sildenafil(inhibitor of cGMP phosphodiesterase): Erectile dysfunction




AMP-Activated Protein Kinase(AMPK)

  • A trimeric serine/threonine kinase composed of a catalytic ? subunit and non-catalytic ? and ? subunits
  • gamma subunit is the AMP receptor(also binds ATP)
  • AMP biding activates AMPK and ATP binding deactivates it
  • this is considered an intracellular energy sensor
  • several upstream kinases can also activate AMPK(LKB1, PLCs, CaMKK?)
  • effects many metabolic pathways to increase ATP production




Huntington’s Chorea

  • progressive motor disability, cognitive and personality decline, and autosomal dominant 
  • onset around 35-50 with death from progression 15-18 yrs later (10% present before 20; usually paternal transmission)
  • patients usually die from aspiration pneumonia due to difficulty swallowing
  • caused by presence of mutated Htt gene on chromosome 4
  • 5-35 CAG repeats in normal alle, ;40 high chance disease
  • shows anticipation(worse with each generation)
  • no effective therapy 
  • widespread neuronal loss in striatum(initially hyperactive)





  • nitrogenous base attached to 1′ carbon of sugar backbone
  • becomes nucleotide when a phosphate is added




Phosphodiester Bond

  • between 5′ carbon and 3 prime hydroxyl group (uses the phosphate group
  • creates a 5′ end (phosphorus group) and a 3′ end (hydroxyl group)




Forces bringing double helix together

  • hydrogen bonding (2 for A-T and 3 for G-C)
  • the third bond makes the bond lenght shorter for g-c
  • base stacking 
  • heavily negative charge




B- form DNA structure


  • rise of 3.4 A per base pair
  • 10 base pairs per turn
  • Major groove and minor groove
  • right handed usually 




Steps of DNA replication in Euk

  • origin of replication identification
  • unwinding of DNA helix
  • formation of the replication fork and synthesis of RNA primer
  • DNA synthesis and elongation
  • removal of RNA, fill in gaps and link together
  • reconstitution of chromatin structure




Proteins and Enzymes involved in 

DNA replication

  • DNA polymerase
  • Helicases: processive unwinding of DNA
  • Topoisomerases: Torsional strain relief
  • DNA primase: initiates synthesis of RNA primers
  • DNA ligase: Seals single strand nicks
  • Single stranded binding proteins: keeps single strands apart during replication





  • cleaves two of the three phosphates and provides link to the 3′ hydroxyl




Autosomal Dominant

  • often seen in multiple generations
  • mothers and fathers are equally likely to transmit or inherit the disorder
  • sons and daughters of an affeted parent are equally likely to inherit and transmit the disorder 




Autosomal Recessive Inheritance

  • horizontal: usually seen in a single generation of a family
  • mothers and fathers are equally likely to transmit or inherit the disorder




Multifactorial Inheritance

  • a single trait/phenotype is controlled by multiple genes and/or environment
  • when plotting array of phenotype you get bellshaped curve (ex is for height:10-15 genes control)
  • high blood pressure and weight are another ex.
  • possible threshold for expression is how many u need to manifest?




Risk estimation for discrete

multifactorial traits

  • risk increases with number of affected relatives
  • risk increases with the severity of the malformation or disease
  • when something occurs more commonly in one gender, then the offspring of the affected individual of the sex thats less common then higher relative risk
  • reccurence risk represents average risks and will vary among different families




pyloric stenosis

  • abnormal thickening of the outlet of the stomach
  • acuses obstruction of gastric emptying and voming
  • 5 times more common in males then in females
  • this means they have a different threshold
  • multifactorial disease




Heritability from twin studies

  • h= (MZ concordance-DZconcordance)/MZ concordance
  • autism is really high: H2=.95 (this means that the MZ twins got it at a high rate compared to the DZ
  • helps with multifactorial things




Types of RNA

  • Messenger RNA (mRNA)
  • ribosomal RNA (rRNA)
  • Transfer RNA (tRNA): central to protein synthesis as adaptors btw mRNA and aa
  • Micro RNA (miRNA): regulate gene expression typically by blocking translation or inducing degradation of target mRNAs
  • Small  nuclear RNA(snRNAs):involved in modifications needed to generate mature mRNA




Nomenclature for describing

sequence variation

  • g. = genomic DNA
  • c.= coding DNA
  • p. = protein




Prokaryotic RNA synthesis

  • single RNA polymerase
  • simpler promoter
  • single factor for initiation 
  • polycistronic mRNA
  • no RNA processing 




Eukaryotic RNA synthesis

  • multiple RNA polymerases
  • complex promoter regions
  • multiple transcription factors required for initiation
  • monocistronic mRNA 
  • extensive RNA processing




Eukaryotic RNA polymerases

  • RNA pol I – for large rRNA
  • RNA pol II – for mRNA and microRNA
  • RNA pol III – tRNA and 5s rRNA




RNA Polymerase I

  • found in nucleolus
  • transcribes the 3 large ribosomal RNAs as one transcript(45S) that is subsequently cleaved: 28S, 18S and 5.8S rRNA
  • rRNA genes are tandemly linked with many copies per genome
  • Pol I synthesizes approximately 70-80% of the RNA in a cell




RNA Polymerase II

  • Found in the nucleus
  • transcribes genes that encode proteins
  • primary transcripts are subsequently produced into messenger RNA
  • accounts for approximately 3-5% of RNA in a cell (accounts for least abbundant RNA)




RNA Polymerase III

  • found in the nucleus
  • transcribes small RNAs, including 5S rRNA and transfer RNA
  • some pol III transcripts are spliced and/or the RNA sequence is edited after synthesis
  • transcribes approximately 17-20% of the RNA in a cell




Eukaryotic promoters 

  • multiple regulatory sequences
  • some have a TATA box
  • other elements too that guide transcription machinary 
  • depends on the gene
  • might be enhancer elements 




Termination sequences

for transcription

  • one sequence causes a hairpin loop
  • second is rho factor that sits on a sequence near end and moves along and when it catches up to RNA polymerase it terminates it 





Polycistronic mRNA

  • found in prokaryotes
  • RNA polymerase transcribes multiple proteins
  • usually called an operon and all the genes trancribe proteins that usually work together
  • an example is the lactose operon 




Assembly of Basic 

Transcription Apparatus

  • TFII D, along with coactivators and TATA binding protein, binds to promoter
  • TFII A and B then recruited
  • RNA polymerase then sits down next to TFII B
  • TFII F, E, and H then bind to the complex




Generation of mature mRNA

  • capping: serves to seal 5′ end of mRNA and decrease rate of degeneration, serves as recognition site for ribosome
  • poly (A) tail
  • intron splicing




Synthesis of Poly-A Tail

  • polyadenylation signal (AAUAAA) upstream from the termination site
  • poly(A) polymerase uses ATP to add a poly (A) tail




Intron Removal

  • U1 binds/ cuts splice donor site (GU)
  • U2 binds lariate aceptor site(middle of intron) (Adenine)
  • U4, 5, 6 join complex and link to U2
  • U 5 identifiies splice acceptor site
  • intron is cut away as a lariat and the two exons are ligated perfectly
  • elements of splicesome complex are reusable and intron lariant is rapidly degraded




Processing of tRNA in nucleus

  • introns are first removed (unique and doesnt involve splicesome
  • then modification of bases and addition of CAA at the 3′ end (site of AA. attachment




Structure of tRNA

  • 3 loops: dihydrouridine(D) loop, Ribothymidine (T) loop (T-C loop), and anticodon loop that recognizes 
  • 70-95 nucleotides in length
  • a.a. attached to the 3′ most base
  • the T-C loop is involved in ribosome binding
  • anticodon base pairs with the mRNA codon (antiparallel)




RISC Complex

  • formed from the dicer complex that is fromed from pre-miRNA after it leaves the nucleus
  • this can either cause mRNA cleavage or translation repression




Wobble hypothesis

  • allows for not perfect match
  • anticodon C binds with codon G 
  • anticodon A binds with codon U
  • anticodon U binds with codon A or G
  • anticodon G binds with codon U or C
  • anticodon I binds with codon U, C, or A




Point Mutation

  • silent: change that specifies the same A.A.
  • missense: change that specifies a different A.A
  • nonsense: change that produces a stop codon




Insertion Mutation

  • causes frameshift or new AA
  • addition of one or more bases
  • ex is CAG inserts Gln(polyQ),  huntington’s disease




Deletion Mutation

  • frameshift or loss of AA
  • loss of one or more bases
  • example is ?-Thalassemia due to frameshift





  • mRNA
  • Aminoacyl-tRNAs and aminoacyl tRNA synthetases
  • Ribosomes – “ribozymes”
  • Eukaryotic initiation factors (eIFs; G-proteins)
  • Elongation factors EF1A and EF2 (G-proteins)
  • Energy in form of ATP and GTP




Formation of Aminoacyl-tRNA’s

  • each AA has a corresponding aminoacyl-tRNA synthetase
  • only 1 tRNA which recognizes the AUG start codon: methionyl-tRNAiMet 
  • charging a tRNA requires energy in the form of ATP and is a two step process




Combing of an AA to tRNA

  • first aminoacyl tRNA synthetase (enzyme) attaches to an amino acid using ATP
  • this creates a Enzyme-aminoacyl-AMP complex
  • it then addes the AA to a tRNA and the AMP and enzyme dissociate
  • atRNA synthetase recognizes three sequences( on d loop, anticodon and 3′ end) in order to attach right AA to right tRNA




Three Steps of Translation

  • Initiation: binding of initiator tRNA to initiator codon at P site of ribosome
  • Elongation: binding of aminoacyl-tRNA to A site; formation of a peptide bond; translocation; ejection of spent tRNA
  • termination: release of completed peptide




Euk. Translation Initiation 


  • capped mRNA
  • small 40S ribosomal subunit
  • eIFs (eIF2 with GTP binds initially to tRNA)
  • initiation Met-tRNA: complex uses ATP to scan the mRNA for Kozak AUG start sites (eIF4F helps here); GTP (on eIF) is then hydrolyzed and eIFs are release
  • large 60S subunit binds to complete the ribosome
  • eIF3 had been stoping large subunit from initially binding




Eukaryotic Initiation of Translation

  • cap at 5′ end of mRNA binds eIF4F and 40S subunit with met-tRNA
  • mRNA scanned for AUG within a kozak consensus sequence
  • 12 or more eIF
  • 80S ribosomes (60 and 40 subunits)
  • First AA is methionine




Prokaryotic Initiation of Translation

  • mRNA has no 5′ cap
  • Shine-Dalgarno sequence upstream of AUG  binds to complementary sequence of 16S rRNA
  • 3 IF
  • 70S ribosome (50S+30S)
  • first AA is formyl-methionine




Translation Elongation

  • eEF1A +GTP + tRNA attach to A site and GTP is hydrolyzed
  • RNA then catalyzes the peptide bond formation(peptidyl transferase)
  • eEF2+GTP then move the tRNA with AA chain from A to P site (translocation)
  • tRNA that was in P site moves to E site and is ejected, along with eEF2 and GTP is hydrolyzed





  • helps in the recycling of eEF1A in translation 
  • eEF1B binds GTP
  • it then exchanges the GTP for the GDP bound to eEF1A after it had went through a cylce of translation
  • makes translation go much faster




Termination of Translation

  • no tRNAs pair with STOP codons
  • protein release factors (RFs) bind to stop codons: RF-1 binds UAA and UAG, and RF-2 binds UAA and UGA
  • binding of RFs cause peptidyltransferase to hydrolyze the peptide-tRNA linkage and the polypetide is release
  • tRNA is released from P site and the two ribosomal subunits separate as a result of GTP hydrolysis




Energy Requirements for Translation

  • multiple high energy bonds required for each peptide bond made: 2 ATP for charging aminoacyl tRNA, 1 GTP is used to bind tRNA to A site, and 1 GTP to translocate tRNA peptide from A to P site
  • Energy requirements at initiation and termination: 2 ATPs for charging Met-tRNA, ATP needed for helicase to undwind hairpins/loops in mRNA structure
  • 1GTP to form initiation complex at AUG
  • 1 GTP for termination associated with dissociation of the ribosomal subunits




Inhibition of Translation- Antibiotics

  • streptomycin: inhibits initiation and causes misreading of the RNA
  • tetracycline: binds small (30S) subunit and inhibits binding of aminoacyl-tRNA
  • chloramphenicol: inhibits peptidyl transferase activity of the large subunit (50S)
  • erythromycin: binds reversibly to large subunit(50S) and inhibits translocation/peptidyl transferase




Targeting sequences in the Polypeptide

  • secretory pathway-N-Terminal signal peptides
  • ER
  • membrane insertion sequences-hydrophobic helices
  • nuclear localization sequences
  • mitochondrial import sequences
  • peroxisome signals




Post translational modifications


  • glycosylation-golgi, lysosomal signals
  • proteolytic cleavage sites
  • phosphorylation
  • lipidation




Core Sugar Complex

  • formed in the lumen of the endoplasmic reticulum 
  • formed from seqential addition of sugar residues at non-reducing end of dolichol phosphate 
  • this core sugar can be added to a protein, and influences the protein folding (hydrophillic)




Mannose Phosphate

  • can be added to an protein
  • mannose-P receptors bind it and a vessicle is formed that transport the protein to the lysosome




Types of DNA binding motif

  • assist in transcription factors bind to DNA
  • Zinc fingers
  • leucine zipper
  • helix-turn-helix
  • helix-loop-helix




Zinc Finger Motif

  • require a chelating agent? (Zinc)
  • must chelate either histidine or cysteine to stabilize the finger motif
  • recognize specifc sequence by grabbing the DNA
  • NRS(alpha helix) recognizes specific DNA sequence on major groove of DNA




Leucine Zipper


  • 7 hydrophobic luecines are alligned on one side of the protein and the other side are polypeptides
  • bond using hydrophobic contraction
  • usually homo or heterodimer transcription factors 
  • arginine and lysine positive amino acid can bind to something in major groove?? 






  • monomer??
  • first helix recognizes DNA major grove
  • 2nd and 3rd stabilize DNA binding
  • first helix is like luecine zipper and has a polypeptide that recognizes major groove using positive charged arginine or lysine 




DNA Response Elements

  • direct repeat: AGGTCAnAGGTCA – called DR1
  • add more “n” to make DR2, DR4, ect
  • palindromes are either inverted(AGAACA(n) TGTTCT) or everted: TGACCT(n) AGGTCA)




Ligand Dependent Transcription factor Activation

(nuclear receptors)

  • LBD; 12 aplha-helices(H1-H12) and 2 beta turns are arranged in three layers
  • a few helices (H3, 5, 10, 11) form a ligand binding pocket that is struturallly conserved hydrophobic surface for ligand interaction
  • H12 in the AF-2 domain hangs out the ligand binding pocket like a waving arm, blocking coactivator recruitment
  • upon ligand binding, conformation chages cause H12 to reposition and allow AF-2 to be functional to recruit coactivators via LxxLL NR box
  • in process, H12 encloses the binding pocket and forms additional H-bond with ligand to stabilize the conformation




Steroid receptors


  • nuclear receptor
  • recognize AGAACA direct sequence
  • found in cytosol 
  • ER recognizes AGGTCA
  • GR(glucocorticoid), MR(mineralcorticoid), PR (progesterone), AR(androgen), ER(estrogen)




RXR heterodimers


  • nuclear receptor
  • AGGTCA is recognition sequence
  • TR(thyroid): DR-4 , VDR(vitamin D3): DR3, PPAR?(fatty acids): DR1, FXR(bile acids): IR1, LXR(oxysterols): DR4, RAR(retinoic acid): DR1
  • TR, VDR and RAR occupy 3′ end and RXR the 5′ end
  • PPAR, FXR recognize 5′ site and RXR at 3′




Class II and III

(thyroid hormone receptor)


  • thyroid hormone receptor always in nucleus 
  • T3 recognizes the TR, this then recruits coactivator
  • when ligand not present, the co-repressor complex containing HDAC will be present




Class I Receptor


  • Glucocorticoid receptor
  • in cytosol and usually associated with HSP(Heat shock protein)
  • when ligand binds, HSP dissociates and GR receptor moves to nucleus




Peroxisome Proliferator Activated Receptor



  • normally expressed in liver
  • invaolved in beta oxdiation
  • ligands are polyunsaturated fatty acid(PUFA)
  • also involved in anti-inflamatory function
  • drug ligands are fibrate drugs, cofibrate, benzafibrate, fenofibrate
  • recognizes DR1: AGGTCA
  • targets acyl CoA synthetase, CPT1, and reduces ApoCIII




Peroxisome Proliferator Activated Receptor



  • ligand is polyunsaturated fatty acid (PUFA)
  • expressed in adipose tissue
  • drugs are TZD avandia
  • target lipoprotein lipase, SCD1, and GLUT4
  • decreases serum glucose and FA, increaes insulin sensitivity, increase number of adipocytes, results in weight gain
  • target for antidiabetic drugs
  • oposite from PPAR alpha




Peroxisome Proliferator Activated Receptor



  • highly expressed in muscle
  • ligands are polyunsaturated fatty acids(PUFA)
  • switch type II(fast and glycolytic) muscle fiber to type I fiber (slow and oxidative phosphorylation)
  • increases endurance with exercise
  • target genes are CPT!, uncoupling protein, lipoprotein lipase




cAMP Responsive Element Binding Protein


  • basic luicine zipper transcription factor that binds to palomdromic sequence as homodimer
  • regulates gluconeogensis
  • Glucagon binds to Gprotein receptor wihc activatys AC which creates cAMP which activates PKA
  • PKA activates CREB activates PEPCK(major gluconeogenic gene)  by recruiting CBP and TORC2 coactivators





  • anti diabetic drug
  • blocks CREB activation(hepatic gluconeogenesis)
  • activates LKB which activates AMPK
  • AMPK phosphorylates TORC2 which causes it to leave the nucleus, where it is supposed to coactivate PEPK with CREB




Foxhead transcription factor

(FOX proteins)


  • involved in insulin signaling
  • winged helix TR that bind as monomer or dimer
  • Insulin normally activates IRS2 which activates AKT which phosphorylates Fox01 which causes it to leave the nucleus
  • Fox01 normally binds to IRE and promotes PEPCK genes
  • FoxA2 regulates FA oxidation and ketogenesis during fasting and it is sensitive to IRS1 and 2
  • insulin resistance decreases IRS2 signaling(hyperinsulin anemia)-Fox1 would be in nucleus and FoxA2 in cytosol




Sterol Responsive Element Binding Protein




  • important for lipid and cholesterol regulation
  • basic helix-loop-helix leucine zipper trans. factor
  • in ER mebrane as associated with SCAP and INSIG(er protein especially)
  • when cholesterol is low they translocate to Golgi where S1P and S2P cleave SREBP and release active form of SREBP-then moves to nucleus and promotes SRE gene transcription
  • SREBP2: cholesterol synthesis and uptake
  • SREB1: fatty acid synthesis: 
  • both found frequently in liver?





  • when DNA is being replicated there is a 3′ overhang on the leading strand. this is a telomeric sequence
  • a telomerase binds here and protects it from being chewed away
  • makes a long telomerase sequence end that wont matter if it is chewed away




DNA Repair Systems


  • must be able to recognize the wrong base or bases
  • must be able to remove the wrong base or bases
  • must repair the newly made gap with a DNA polymerase and DNA ligase activity
  • mismatch repair(copy errors, single base or up to 5), base excision repair(env. chemical radiation, spontaneous degradation), nucleotide excision repair(chemical or radiation damage to a DNA segment), double strand break repair(ionizing radiation, chemotherapy, oxidative free radicals)




Werner Syndrome

  • is the gene on the 8th chromosome that codes for a DNA helicase involved in many DNA repair and processing pathways
  • helicase may also have a role in telomere maintenance
  • no known conventional therapy for this genetic defect
  • autosomal recessive 





Regulation of operons by repression

  • operon normally inhibited by a represser at the operator sequence upstream from the structural genes
  • an inducer (lactose) can come in and bind with the repressor, thus inactivating it 
  • this allows for RNA polymerase to bind




Lactose Operon

  • allolactose binds to repressor, inhibiting it
  • in presence of lactose and glucose, the genes are on but not transcribing very fast
  • when glucose is low and theres lactose, cAMP activates CRP that binds to the promoter
  • this speeds up transcription 




Eukaryotic Gene Expression


  • initiation of transcription
  • RNA processing(cap and poly A addition)
  • RNA splicing and alternative splicing
  • transport of mRNA from the nucleus to the cytoplasm
  • regulation of mRNA stability-microRNA(miRNA)
  • gene amplification
  • gene rearrangement 




Chromatin Remodeling

  • modifiers such as acetyltransferases can promote or deter the activity of more substantial chromatin remodelers
  • chromatin remodelers organize the nucleosomes to promote gene activation or repression
  • such remodelers may either slide or eject necleosomes to promote DNA access to other transcription factors
  • it transfers a neutral acetyl group to a positive N of lysine, this causes the bond between histone protein and DNA to lessen because it was postive to the DNA negative before




Phases of the Cell Cycle

  • G1: 6-12 hours
  • S phase(DNA replication): 6-8 hours
  • G2: 3-4 hours
  • M (mitosis): 1 hour




Cyclin binding

  • cyclin binds to CDK, moving the blocking group
  • this allows a threonine to become phosphorylated 
  • this then activates the CDK




CDKs and the cell cycle

  • G1 up to restriction point is coordinated by CDK4/6 that is activated by Cyclin D (allows cell to go past restriction point)
  • After restiction point CDK2 and Cyclin E push the cell into S phase
  • CDK2 and cyclin A then push the cell through S phase to G2 phase
  • CDK1 and cyclin A/B pushes cell through G2 phase
  • CDK1 and cyclin A/B also push cell through mitosis




Cell Cycle Check Points

  • surveillance points which help ensure the mother cell passes accurate copies of its genome to the two daughter cells
  • is the cell ready for division?, does the replicated genome contain errors, or is damaged?, Is DNA synthesis complete?, Are chromatids properly assembled on mitotic spindle? 




Stop Sign/Restriction Point

  • first a growth factor initiations a cellular pathway(Ras/Raf)
  • eventually Cyclin D is produced and it binds to CDK4
  • this phosphorylates retinol blastoma protein(Rb) which is a tumor supressor and inhibitor
  • this inactivates it, freeing up E2F
  • E2F then activates a massive series of genes to progess the cell cycle
  • CKI can block the CDKs from working
  • Cylin E and CDK2 can also help in inactiving the Rb




CKI’s Cyclin Kinase Inhibitors

  • two classes: those that inhibit CDKs 1, 2, 4, 6 (p21) and those that only inhibit CDK 4 and CDK 6
  • these are proteins that directly bind to the cyclin-CDK compleses to inhibit CDK activity
  • p21 has an important role in stopping the cell cycle when DNA damage occurs





  • checkpoint protein(tumors can result from mutation)
  • when it is induced: 
  • signals that theres damage in DNA, so it induces p21
  • if it doesnt get fixed, it induces apoptosis (activates caspases)




Steps of Mitosis

  • prophase
  • prometaphase
  • metaphase
  • anaphase
  • telophase




Mitosis Simplified


  • abrubt increas in M-CDK(cyclin B) activity drives early mitosis events(chromosome condensation and alignment
  • active CDK1 activates proteins that lead to the assembly of mitotoic spindles and chromatid attachment




Mitosis Simplified


  • anaphase promoting comples(APC/C) helps drive separation of the sister chromatids
  • anaphase promoting complex(APC/C) degrades cyclins also degrades other anaphase inhibitory proteins




Major Classes of Signals

for Eukaryotic Cell Growth

  • mitogens(activate cell cycle)
  • cell growth factors (increase cell size, promote protein synthesis)
  • survival factors (mainly inhibitors of apoptosis)
  • all three of these are operating in cancer cells





  • used for transplants because it is an immunosupresnet
  • inhibits mTOR?






basics of activation

  • integrator of nutrients signals to cell growth
  • rampamycin, mitogens, amino acids, energy and DNA damage effect it to do other thing
  • must be activated during process of growth
  • serine-threonine kinase
  • after activated it finds targets to phosphorylate 
  • some of its effects are cell migration, cell cycle progression, cell growth/size, cell survival, autophagy
  • can be activated by a G-protein that is dependent on low levels of AMP
  • GPrtotein also activated by AKT that is activated by insulin
  • mTOR activation signals to allow ribosome translation***






  • TOR enzyme complex is a central controller for cell growth(accumulation of mass
  • metazoan(multi tissue) TOR is essential for growth during early development(deletion of the TOR gene is embryonic lethal in mammals)
  • TOR inclunce in mature or aging organisms is partly negative: unwanted cell proliferation; continued growth without division may lead to protein aggregates and abnormal proteins that may induce late life malfunction





  • activated by mTORC1
  • this then can then activate lipid biogenesis, metabolism, translation, cell size regulation, DNA replication, RNA processing
  • also takes part in a negative feedback mech by inhibiting insulin receptor substrate
  • mTORC1 can also activate Grb10 that inhibits the insulin like growth factor receptor




Medical Genetics

  • among the 5000 listed genetic diseases, over 1100 genes can be tested clinically
  • half of the 2% of fetuses with major genetic abnormalities have abnormal chromosomes
  • 30% of children admitted to the hosphital have a genetically related problems




Centromere Location

  • metacentric: when centromere is in the center
  • submetacentric: when centromere is closer to one side then the other (but both still the same length)
  • acrocentric: when the centromere is at one end and one is sticks out a little farther then the other one
  • p arm is the short arm, q arm is the long arm




Banding patterns

  • can depend on different ratios of the basepairs
  • if stained with an A-T binding protein then parts of a chromosome would look brighter if it had A-T and dimmer if it had G-C
  • when fluorescing basepairs, the more in a row, the more exponentially bright it is(2 – 4x brightness, 3- 9x brightness)




Acrocentric chromosomes

  • contain stalks and satellites
  • about 10 chromosomes like this
  • ribosomal genes tend to be on stalks




Down Syndrome from Trisomy 21

  • palpebral fissures are slanted upward and outward
  • flat nasal bridge
  • round face
  • small mouth, thick lips, often large tongue
  • average IQ of 50 at 5 years, 38 at 15 years and later degenrates further
  • hypotonic infants (floppy)




Nondirective counseling

  • provides all known information and allows the patient to decide
  • couple informed of fetal status, advised of the possibilities, and then advised to decide without the help of a counselor or physician but with trusted individuals and families with similarly affected patients




Molecular Cytogenetics

  • can test chromosome copy number and single gene copy number
  • enabled by cloned genes and other DNA fragments that can be sequenced, labeled with specific fluorescent colors, and hydridized specifically to similar or identical DNA sequences on chromosome bands, and viewed at 1000-fold magnification




Microarray Principles

  • microarrays test hundres to many thousands of specific chromosome targets simultaneously by labeling total DNA from a patient sample in the first color and total DNA from normal DNA in the second color and hybridizing these to all of the specific cloned genomic sites bound to previously designated locations on the microarray
  • comparison of both fluorescent colored signals at each location determines whether the patients DNA has less, the same, or more copies of the DNA target than normal DNA. many locations to small to be seen on banding or FISH can be tested at the same time to look for abnormal gene copy numbers in doezens of gene disorders as well as abnormal chromosome locations
  • FISH is the clinically validated test used to confirm abnormal microarray results





  • may be inherited or arise following two gene mutations to both retinoblastoma genes in the same cell
  • used to pluck out the eye
  • looks like white spot in eye
  • prenatal diagnosis, routine eye exam
  • laser beam destroys early tumors
  • inherit one bad gene, and mutation causes other, or two bad mutations 




Other approaches to 

Treating genetic diseases

  • 3. transplantation(adult polycystic kidney disease)
  • 4. modifying diet (maternal PKU)
  • 5. Gene therapy(adenosine deaminase)




Enabling PCR Attributes

  • primer selection: four different basepair possibilities at each of 17 locations
  • temperature resistant polymerase from bacteria growing in hot springs allows repeated cycling
  • logarithmic amplification of target sequence




Chorionic Villus Sampling(CVS)/ Maternal Decidual Contamination

  • used to test placenta 
  • tests for circulating fetal DNA in maternal circulation
  • one type of error though is there could be abnormal cells in circulation but the fetus is actually normal





  • occurs after fertilization at the 64-128 cell stage of the blastocyst(embryo) and inactivates about 80% of the genes
  • results in turning off the genes during the remainder of the life cycle prior to gametogenesis or fertilization. Methylation of the 5′ (upstream) promoter region of the genes is involved
  • the XIST(Xinactivation gene) in proximal long arm initiates inactivation 
  • turns off about 70% of the genes on average




Perutz Nobel Prize 1962

  • hemoglobin protein conformation




Gene Deletion (decreased)


  • PMP22: hereditary neuropathy with pressure palsy
  • SNRPN: Prader-Willi Syndrome
  • UBE3A: Angelman Syndrome (adjacent to SNRPN)




Gene Duplication(increase)


  • PMP22: Charcot-MArie-Tooth Type IA
  • SNRPN: Autism 




Imprinting and Uniparental Disomy(decearsed)


  • SNRPN: Prader-Willi Syndrome
  • UBE3A: Angelman Syndrom (adjacent to SNRPN)




Diagnostic Criteria for Autism

  • severe abnormality of reciprocal social relatedness(plays alone, dislikes being held)
  • severe abnormality of communication skills(incl. laguage)
  • restricted, repetitive behavior, interests, activities, and imagination
  • early onset (before age 5)
  • can be caused by extra genes upstream?




UPD-Uniparental Disomy

  • starts out with three copies of a chromosome at fertilization
  • one needs to be lost: if the parent that gave two’s chromosome is lost it would be a normal individual
  • If one from the father is lost then two from a single parent (1/15000)
  • gene turning off??




Reported Abnormal Uniparental Disomy

  • Parental and Maternal: Chromosomes 14 and 15
  • Paternal: Chromosomes 1, 2, 6, 7, 11(lethal)
  • Maternal: chromosomes 9, 16, 20, 22
  • Suggested: chromosomes 4, 5, 13, 21, X




Micro Array

  • used to define deletions and duplications of large regions of a genome
  • use chips with attached segments of genetic material
  • have test DNA and reference DNA
  • if they both bind to same site, mix of color
  • ratio of test dna/reference is elevated then evidence of duplication
  • if ratio is under represented then evidence of deletion




Classic Sanger Sequencing


  • took segment of DNA and chopped up into parts with nucleases
  • PCR amplified it
  • add in nucletides (some labled didioxynucletides)
  • created random length segments and sized them on gel
  • in direction of larger size you can read the sequence
  • would take a long time
  • new method is similar to the old but uses computers and lasers to sequence after multiplying and matching fragments in parallel





Allelic Heterozygosity

  • different mutations in same locus
  • a disease can be more or less severe depending on where in the allele the  mutation happens
  • severity caussd by the difference in the particular alleles




Locus Heterozygosity

  • multiple genes actually contribute to same phenotypic trait
  • classic example is retinitis pignitosum: can be due to any one of 15 gene mutations (several autosomal dominant, recessive and x linked) 




Variable Expression within a family

  • both mother and son can have an autosomal dominant defect but mother is much less severe
  • this could be due to modifying genes-one or more independent gene or sequence involved in the expression of a phenotype





  • extreme example of variable expression
  • gene is present but not manifest
  • modifying genes work in such a magnitude that you cant even detect the problem in the one not manifesting it
  • equation is number of mutations with phenotype divided by number of individuals with mutation x 100
  • complete penetrance is when it = 100% and incomplete is anything less




Delayed Onset Dominant Conditions

  • example is poly cystic kidney disease
  • results from one heritable mutation and one somatic mutation
  • the somatic mutation in ADPKD would be inside kidney cystic cells and the autosomal would be in all cells





  • matters which parent passes on a mutation in an imprinted gene
  • if the gene is inactivated in the mother and active in the father-then it will only be manifest if the mutant gene is inherited from the father
  • imprinted is erased and re-established with the production of gametes




Considerations about use 

of Recombinant DNA

  • must also include appropriate promotor when inserting gene
  • if bacterial system used, the inserted DNA has to be of viral or bacterial orgigin or the cDNA sequence from eukaryotic genes(no introns)
  • yeast have advantage of being euk. so can process pro-mRNA
  • simple peptide products are made. if the peptide requires further modification, it likely has to be performed separately
  • in most cases the producing cell has to be disrupted and the product purified




Products produced by recombinant 

DNA technology

  • vaccines-hep B
  • peptide hormones-insulin, prathyroid hormone, growth hormone
  • bone marrow stimulating factors-erythropoietin, GM-CSF, G-CSF
  • coagulation factors-factor VIII, factor IX and tissue plasminogen activator
  • interferons and several interlukins




Treatment of Human Genetic Disease

  • symptomatic treatement is most common
  • replacement therapy
  • compensation for functional deficits using drugs
  • use of small molecules as salvage agents
  • transplantation (provide stem cells from normal donor, risk of conditioning treatment and GVHD)
  • DNA transfer therapy(gene therapy)





  • used in a number of cases: bone marrow for Hg SS, Liver for some MPS, kidney for familial FSGS, polycystic kidney
  • issue of ability to find transplant and risks of longstanding immunosuppression
  • natural extension would be to transplant manipulated stem cells derived from patient




Issues with gene therapy

  • delivery of the material and place of delivery(transfection, direct insertion
  • sustained expression – stem cell vs somatic cell
  • avoidance of deleterious immunologic or tissue based responses
  • potential for inducing secondary disease based on DNA disruption by DNA insertion




Epigenetics (modern definition)

  • modifications of DNA or associated proteins that are not base pair sequence variations that carry information from mother cell to daughter cells
  • these modifications result in a heritable phenotype without changes in the DNA sequence




types of epigenetic information

  • modification of chromatin structure (CTCF)
  • chromatin factors(trithorax proteins, polycomb group proteins)
  • histone modification: methylation, acetylations, ubiquitinylation, and phosphorylation
  • methylation of DNA 





  • DNA methyltransferases
  • recognize the CpG in DNA and then transfers methyl group to it
  • series of enzymes that can later remove this methylation (TETs)




Imprinted Genes

  • methylated genes that keep there methylation during fertilization
  • fair amount on fathers genes and less so on mothers
  • imprinted dont change during genome wide demethylation (De nevo genome wide methyl.)
  • homologs from mother are expressing so only hers effect body when fathers are imprinted
  • violates usual rule of inheritance(usually both alleles in a heterozygote are equally expressed)
  •  cells that are destined to make gamates lack all imprinting




Hyper vs Hypomethylation Responses

  • if one is hypermethylated, they would have a lack of response to an environmental stimulus
  • if they are hypomethylated they would have a heightened response to the same stimulus
  • cancer tends to come from cyclin dependent kinase inhibitor (CKI’s) genes being hypermethylated




Genomic Imprinting Disorders

  • Prader-Willi syndrome (fathers gene is mutated)
  • Angelman syndrome (mothers gene is mutated)
  • both on chromosome 15qII
  • mothers gene is shut off already in prader
  • fathers gene shut off already in angelman




Prader-Willi Syndrome


  • Usually caused by micro deletion in region q11-13 of the paternally transmitted chromosome 15
  • several genes in this region are genomically imprinted in the maternal chromosome(turned off)
  • hence, if there is a paternal deletion in this region, there are no active genes and you get the syndrome
  • mental retardation, hypotonia, uncontrolable appetite, paternal chromosome has deletion




Angelman Syndrom


  • Usually caused by micro deletion in region q11-13 of the maternally transmitted chromosome 15
  • several genes in this region are genomically imprinted in the paternal chromosome(turned off)
  • hence, if there is a maternal deletion in this region, there are no active genes and you get the syndrome
  • microcephaly, severe mental retardation with no speech, and jerky ataxic movements; maternal chromosone has a deletion





  • produces a critical ubiquitin ligase enzyme
  • requires the maternal allele to be expressed in brain during development
  • Angelman syndrom involves a deletion of this gene in the maternal genome




Genetic Testing for Medical Care

  • establish the diagnosis in a symptomatic person
  • predictive testing for a relative at risk for a treatable disorder (early treatment, surveilance for a treatable manifestations that appear over time)




Genetic Testing For Personal 


  • predictive testing for a relative at risk for an untreatable disorder
  • carrier testing(autosomal recessive or x linked recessive)
  • prenatal diagnosis/preimplantation genetic diagnosis (PGD)




Phenotype First Approach

(genetic testing)

  • clinician uses detailed phenotypic findings to determine gene(s) most likely to be mutated
  • clinician selects test method(s) most likely to detect a mutation 
  • mutation screen including most common known mutations, sequence analysis, deletion/duplication testing(MPLA, qPCR)




Spinal Cerebellar Atrophy, type 3


  • autosomal dominant disease that results from expansion of a CAG trinucleotide in the ATXN3 gene
  • 100% of cases can be detected by assessing the CAG Repeat number
  • ataxic gait, sensory deficit, speech problems, tongue fasiculations, pyramidal signs




Tay-Sach’s Disease

  • autosomal recessive condition
  • neuronal storage disease
  • a mutation of HEX A gene resulting in nonfunctional hexosaminidase A
  • six mutations that have a 98% chance of being detected




Bayes Theorem

  • multiple independent determinations of risk can be combined to result ina  much refined risk extimate
  • start with the prior(baseline) probability
  • multiply this with conditional probabilites determined by biochemical or genetic testing-arrive at joint prob.
  • can then determine a posterior(final) prob by dividing the joint prob. of the condition of interest by the sum of all joint prob. 




Presymptomatic testing

  • testing where if you find a mutation, you will develop the disease
  • an example is huntingtons disease
  • if you dont have the mutation, you will not develop symptoms of the disorder
  • issues include effect of knowing that the condition will develop and consequences for employment and health insurance




Predispositional Testing

  • if mutation is found you have a higher risk of developing the disorder but not 100%
  • if the mutation is not found you have a lower risk of developing the disorder but not 0%
  • issues are ability to modify outcome, stress of uncertainty and possible discrimination




Genotype First Aproach

(genetic testing)

  • often used for broad phenotypes(ex develop delay, multiple anomalies)
  • use multi-gene panals (genes included and test methods vary by lab)
  • whole exome sequencing (WES): sequencing of all coding regions = ? 1% of genome
  • CGH based array methods




Problems with Genotype First

Genetic Testiing

  • may not find anything, what is detection rate?
  • more genes tested, more variants of unkown significance
  • patients phenotype may not fit well with what is known about the disorder at the time




Prenatal Diagnosis

  • sample-parental samples, fetal DNA in maternal serum, chorionic villus sampling or amniocentesis, IVF with blastomere biopsies
  • Testing-FISH and/or karytype, limited or whole genome array CGH: single gene, gene panal or exome sequencing
  • issues: timing(only 20wks from finding out), significance of results, collateral effects(ultrasound might detect structurally abnormal fetus), cost

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