Biochemistry – Lecture 25

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X occur widely as components of antibiotics (derived from various sources)
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oligosaccharides
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How is erythromycin produced?
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is an antibiotic produced by a strain of Streptomyces erythreus
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is an antibiotic produced by a strain of Streptomyces erythreus
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erythromycin
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***Erythromycin – It’s an important antibiotic 1. What does it do and how? 2. Compared to penicillin? 3. Side effects?
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1. Erthryomycin binds to the 50s subunit of the bacterial 70s rRNA complex, and protein synthesis and subsequently structure/function processes critical for life or replication are inhibited. 2. Erythromycin is an antibiotic that has an antimicrobial spectrum similar to or slightly wider than that of penicillin, and is often used for people who have an allergy to penicillins. For respiratory tract infections, it has better coverage of atypical organisms, including mycoplasma (a genus of bacteria without a cell wall) and Legionellosis (stains poorly with Gram stain because of unique lipopolysaccharide coat, requires silver staining to visualize) 3. Gastrointestinal disturbances, such as diarrhea, nausea, abdominal pain, and vomiting are very common
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binds to the 50s subunit of the bacterial 70s rRNA complex, and protein synthesis and subsequently structure/function processes critical for life or replication are inhibited.
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Erythromycin
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is an antibiotic that has an antimicrobial spectrum similar to or slightly wider than that of penicillin, and is often used for people who have an allergy to penicillins. For respiratory tract infections, it has better coverage of atypical organisms, including mycoplasma (a genus of bacteria without a cell wall) and Legionellosis (stains poorly with Gram stain because of unique lipopolysaccharide coat, requires silver staining to visualize)
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Erythromycin
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Erthryomycin binds to the x of the bacterial x, and protein synthesis and subsequently structure/function processes critical for life or replication are inhibited.
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50s subunit of the bacterial 70s rRNA complex
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Mycoplasma
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a genus of bacteria without a cell wall
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Stains poorly with Gram-stain because of unique lipopolysaccharide coat, requires silver stain to visualize
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Legionellosis
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How to stain for Legionella
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It’s a gram negative bacteria, and you need to stain with silver to visualize it.
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Legionnaires’ disease is caused by
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Legionella bacteria
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Legionellosis 1. What is it? 2. History 3. When was it identified and isolated? 4. How do you visualize it?
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1. It’s a disease caused by Legionellosis bacteria that causes illness, pneumonia, rapid breathing then chest pain. Can cause death 2. First recognized outbreak occurred on July 27, 1976 at the Bellevue Stratford Hotel in Philadelphia, where members of the American Legion, a United States military veterans association, had gathered for the American bicentennial. Within two days of the event’s start, veterans began falling ill with a then unidentified pneumonia. They had rapid breathing and complained of chest pain. As many as 221 people were given medical treatment, and 34 deaths occurred. At the time, the U.S. was debating the risk of a possible swine flu epidemic, and this incident prompted the passage of a national swine flu vaccination program. That cause was ruled out, and research continued for months, and various theories proposed – from toxic chemicals to terrorism aimed at the veterans. U.S CDCP mounted an unprecedented investigation and by September ended up focusing on the hotel environment itself and found in January 1977 the true cause, the Legionellosis bacterium. It was finally identified and isolated, and was found to be breeding in the cooling tower of the hotel’s air conditioning system, which then spread it through the entire building. This prompted new regulations worldwide for climate control systems. 3. Identified in January 1977. 4. Have to do a silver stain
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Vancomycin 1. What is it? 2. How does it act in general terms? 3. How does it act specifically? 4. Describe the chemistry and molecules involved that vancomycin binds
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1. is a glycopeptide antibiotic used in the treatment of infections caused by Gram positive bacteria 2. Vancomycin acts by inhibiting proper cell wall synthesis in GRAM POSITIVE bacteria. Due to different mechanism by which gram negative bacteria produce their cell walls and the various factors related to entering the outer membrane of Gram-negative organisms. Vancomycin is not active against Gram-negative bacteria. 3. Vancomycin prevents incorporation of N-acetylmuramic acid (NAM) and N-acetylglucosamine peptide subunits into the peptidoglycan matrix; which forms the major structural component of Gram-positive cell walls. Basically, it ruins the base of everything 4. Vancomycin is a ****ing huge hydrophilic molecule that’s able to form hydrogen bond interactions with the terminal D-alanyl-D-alanine moieties of the NAM/NAG peptides. This binding of vancomycin to the D-alanyl-D-alanine prevents the incorporation of the NAM/NAG subunits into the peptidoglycan matrix.
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When is Vancomycin used?
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Vancomycin has traditionally been reserved as a drug of “last resort”, used only after treatment with other antibiotics had failed, although the emergence of vancomycin resistant organisms means that it is increasingly being displaced from this role by new antibiotics. Vancomycin never became first-line treatment for Staphylococcus aureus for several reasons… in other slides
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Does vancomycin work against gram positive or gram negative bacteria? Why?
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Vancomycin acts by inhibiting proper cell wall synthesis in GRAM POSITIVE bacteria. Due to different mechanism by which gram negative bacteria produce their cell walls and the various factors related to entering the outer membrane of Gram-negative organisms. Vancomycin is not active against Gram-negative bacteria.
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How does Vancomycin prevent NAM/NAG-peptide subunit incorporation into the peptidoglycan matrix?
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Vancomycin is a ****ing huge hydrophilic molecule that’s able to form hydrogen bond interactions with the terminal D-alanyl-D-alanine moieties of the NAM/NAG peptides. This binding of vancomycin to the D-alanyl-D-alanine prevents the incorporation of the NAM/NAG subunits into the peptidoglycan matrix.
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has traditionally been reserved as a drug of “last resort”, used only after treatment with other antibiotics had failed, although the emergence of x resistant organisms means that it is increasingly being displaced from this role by new antibiotics. x never became first-line treatment for Staphylococcus aureus for several reasons… in other slides
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Vancomycin
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Why is vancomycin not a first line of treatment for bacterial infections, especially for Staphylococcus aureus?
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1. Poor oral bioavailability: not well absorbed into GI tract, have to give intravenously. 2. Other orally available antibiotics including methicillin (and its successors, nafcillin and cloxacillin) have better activity against staph aureus. Vancomycin is still used for methicillin resistant staph aureus.
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What antibiotics kick the crap out of vancomycin in terms of killing staph aureus?
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methicillin, nafcillin, cloxacillin
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Dextrans
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It’s a cooler version of starch and glycogen, where the main linkages aren’t alpha(1,4), but they are alpha(1,6). This isn’t amylopectin, where 1,4 is main linkage with occasional 1,6 here and there. No, dextrans are linked 1,6 and have occasional 1,4 1,3 and 1,2 linking adjacent strands
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If you change main glucose linkage from alpha 1,4 to alpha 1,6, you get a new family of polysaccharides – called?
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dextrans
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Roles of dextrans
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1. Dextrans formed by bacteria are components of dental plaque 2. Dextrans in plaque presumably provide protection for oral bacteria 3. Cross linked dextrans are used as “Sephadex” gels in column chromatography 4. These gels, used to separate biomolecules on the basis of size, are up to 98% water.
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X formed by bacteria are components of dental plaque
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dextrans
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dextrans in plaque presumably provide
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protection for oral bacteria
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x are used as “Sephadex” gels in column chromatography
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cross-linked dextrans
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These gels, used to separate biomolecules on the basis of size are up to 98% water
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Sephadex gels made from cross linked dextrans.
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Agarose, a component of agar obtained from marine red algae, is a chain of
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alternating D-galactose and 3,6-anhydro-L-galactose
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used in laboratories to separate biomolecules on the basis of size
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agarose gels (sephadex another answer)
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The favored conformation of agarose in water is…..why does this matter?
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The tertiary structure of agarose is a double helix with a so-called threefold screw axis. The central cavity of this double helix can accommodate H2O molecules. Agarose and agaropectin readily form gels that contain high amounts of H2O (up to 99.5%).
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x are linear chains of repeating disaccharides in which one unit is an amino sugar and one or both is negatively charged.
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Glycosaminoglycans
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Glycosaminoglycans are components of
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proteoglycans
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Glycosaminoglycans
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linear chains of repeating disaccharides in which one unit is an amino sugar and one or both is negatively charged.
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Linear chain of repeating disaccharides
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glycosaminoglycans
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Functions of glycosaminoglycans 1. What is a GAG? 2. Heparin 3. Hyaluronates 3. Chondroitins and keratan sulfate 4. Dermatan sulfate 5. Constituents of?
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1. Linear chains of repeating disaccharides in which one unit is an amino sugar and one or both is negatively charged 2. Heparin, very high negative charge, is natural anticoagulant. 3. Hyaluronates (up to 25,000 disaccharides units) are components of the vitreous humor of the eye and of synovial fluid, the lubricant fluid of the body’s joints. 4. Chondroitins and keratan sulfate are found in tendons, cartilage, and other connective tissue 5. Dermatan sulfate is a component of the extracellular matrix of skin 6. Glycosaminoglycans are constituents of proteoglycans
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Glycosaminoglycan with very high negative charge, a natural anticoagulant.
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heparin
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Hyaluronates
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Glycosaminoglycan structure consisting of up to 25,00 disaccharide units. Component of the vitreous humor of the eye and of synovial fluid, the lubricant fluid of the body’s joints.
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Glycosaminoglycan structure consisting of up to 25,00 disaccharide units. Component of the vitreous humor of the eye and of synovial fluid, the lubricant fluid of the body’s joints.
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Hyaluronates
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Chondroitins and keratan sulfate
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glycosaminoglycan structure found in tendons, cartilage, and other connective tissue
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glycosaminoglycan structure found in tendons, cartilage, and other connective tissue
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Chondroitins and keratan sulfate
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Dermatan sulfate
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glycosaminoglycan component of the extracellular matrix of skin.
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glycosaminoglycan component of the extracellular matrix of skin.
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dermatan sulfate
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Glycosaminoglycans are constituents of
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Proteoglycans, heparin, hyaluronates, chondroitins, keratan sulfate, dermatan sulfate
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Where do you see a huge variety of glycoproteins and proteoglycans
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animal cell surfaces
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***Describe basic proteoglycan structure
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You have GAG units with tetrapeptide bridges?
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Function of cell surface polysaccharides. How do they even show info?
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These polysaccharide structures, glycoproteins and proteoglycans, regulate cell-cell recognition and interaction. The uniqueness of the “information” in these structures is determined by the enzymes that synthesize these polysaccharides
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The uniqueness of the “information” in these structures is determined by the enzymes that synthesize these polysaccharides
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Cell surface polysaccharides/glycoproteins/proteoglycans
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How do proteoglycans modulate processes in cells and organisms?
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Proteoglycans are glycoproteins whose carbohydrates are mostly glycosaminoglycans. Proteoglycans are components of the cell membrane and the glycocalyx – the sugar shell around the cell. Proteoglycans typically consist of proteins with one or two types of glycosaminoglycans
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*****What’s the difference between proteoglycans and glycoproteins?
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Proteoglycans are glycoproteins whose carbohydrates are mostly glycosaminoglycans.
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Where do you find proteoglycans?
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They are components of the cell membrane and the GLYCOCALYX – “sugar shell” around the cell.
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Glycocalyx
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Proteoglycan “sugar shell” around the cell
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Proteoglycan “sugar shell” around the cell
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glycocalyx
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They typically consist of proteins with one or two types of glycosaminoglycan
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proteoglycans
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How is the structure of cartilage matrix proteoglycan important for function?
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The cartilage matrix proteoglycan is highly hydrated. Compression of cartilage (as in walking) squeezes water out of the cartilage tissue to cushion the joint. Water is reabsorbed when the stress and compression diminishes.
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2 major functions of Proteoglycans and examples
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1. Modulation of cell growth processes – proteoglycans bind growth factor proteins in the glycocalyx, providing a reservoir of growth factors at the cell surface 2. Cushioning in joints – Cartilage matrix proteoglycans absorb large amounts of water. During joint movement, cartilage is compressed, expelling water.
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Cartilage matrix proteoglycans absorb large amounts of water. During joint movement, cartilage is x, expelling water.
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compressed
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x bind growth factor proteins in the glycocalyx, providing a reservoir of growth factors at the cell surface
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proteoglycans
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2 big functions of heparin
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Glycosaminoglycan structure with very high negative charge, – Natural anticoagulant – inhibits cell proliferation in a process involving internalization of the GAG moiety and its migration to cell nucleus.
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Heparin – a x glycosaminoglycan, is widely used as an x anticoagulant
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highly sulfated, injectable
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A highly sulfated glycosaminoglycan, widely used as an injectable anticoagulant
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Heparin
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Can be used to form an inner anticoagulant surface on various experimental and medical devices such as test tubes and renal dialysis machines
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heparin
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Heparin acts as an x, preventing the formation of clots and extension of existing clots within the blood. While heparin does not x, it allows the body’s natural clot lysis mechanism to…x
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anticoagulant, does not break down clots that have already formed, it allows body’s natural clot lysis mechanisms to work normally to break down clots that have formed.
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***The Heparin anticoagulation pathway
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Heparin binds to enzyme inhibitor antithrombin III(AT), this causes a conformation change that activates AT through an increase in the flexibility of its reactive site loop. The activated AT then inactivates thrombin and other proteases involved in blood clotting. So, basically Heparin binds and activates AT, and AT inactivates thrombin and other proteases involved in blood clotting.
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Heparin binds and activates
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Enzyme inhibitor antithrombin III (AT)
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Enzyme inhibitor antithrombin III(AT) is activated by, and inactivates…
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activated by heparin, and inactivates thrombin and other proteases involved in blood clotting.
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Proteins that bind carbohydrates with high specificity and high affinity
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Lectins
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Lectins
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proteins that bind carbohydrates with high specificity and high affinity.
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What is the sugar code?
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Sugars are the “letters” of the sugar code. Lectins are “translators” of the sugar code. Many processes, such as cell migration, cell-cell interactions, immune responses, and blood clotting, depend on information transfer using this code.
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Many processes, such as cell migration, cell-cell interactions, immune responses, and blood clotting, depend on information transfer using this code.
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the sugar code – sugars are the letters, lectins are the translators.
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C-Reactive Protein 1. What is it? 2. What does it do? 3. How does it act? 4. Diagnostic marker of x?
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1. A protein (lectin) that is found in the blood, the levels of which rise in response to inflammation. 2. C-reactive protein (CRP) functions to limit tissue damage, acute inflammation, and autoimmune reactions 3. CRP acts by binding to phosphocholine moieties expressed on the surface of dead or dying cells (and some types of bacteria) in order to activate the complement system, a system that helps clear pathogens from the body. 4 CRP is a diagnostic marker of inflammation, and as such it is an indicator of heart disease risk. Since many things can cause elevated CRP, it’s not a specific prognostic indicator.
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A protein found in the blood, the levels of which rise in response to inflammation. Its physiological role is to bind to phosphocholine expressed on the surface of dead or dying cells (and some types of bacteria) in order to activate the complement system, a system that helps clear pathogens from the body.
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C-reactive protein (CRP)
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How does CRP attempt to limit inflammation?
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CRP levels rise dramatically during inflammatory processes occurring in the body. CRP binds to phosphocholine on FOREIGN CELLS/MICROBES and DEAD/DAMAGED CELLS and is thought to assist in complement binding by enhancing phagocytosis by macrophages, which express a receptor for CRP.
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Activity of these goes up when CRP binds to foreign or damaged cells
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Macrophages.
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Express a receptor for CRP
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macrophages – target CRP bound to foreign/dead stuff
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This rises up to 50,000 fold in acute inflammation, such as infection
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CRP
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CRP rises up to x fold in acute inflammation, such as infection
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50,000
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Marker of inflammation
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CRP
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Measuring and charting CRP values can prove useful in
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determining disease progress or the effectiveness of treatments

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