Term
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Definition
1)Massive quantities of mevalonic acid, a percursor of cholesterol was detected in urine.
2)Activity of mevalonate kinase (MKase) was severely deficient in parent's fibroblast and was about HALF-NORMAL IN EACH PARENT.
3)G-->A transversion A334T
Normal Km for aTP BUT Km for mevalonate was increased >50-fold under standard assay condition.
4)A334 is INVARIANT in MKase from bacteria to man (conserved and important for enzyme to do its job).
5)Negative feedback loop for cholesterol
CONCLUSION:
Built of MVA due to impairment of MVAse (Kcat deficiency)-less cholesterol&hence induced HMGR activity, Km increase leads to higher levels of MVA needed. |
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Term
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Definition
1)initial velocity vo of rxn is determined from slope of the progress curve at the beginning of the rxn, vo INCREASES with INCREASING S concentration but reaches a limiting value characteristic of each enzyme, velocity at any time is vt.
2)Vo doubles as enzyme concentration doubles.
3)velocity of enzyme rxn is FIRST ORDER with respect to E BUT NOT so with [substrate] |
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Term
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Definition
| in vo vs. [S] curve, we don't have time course, but we see a parabolic curve that approaches vmax as [s] increases (first order, mixed order, than zero order near Vmax asymptote). |
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Term
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Definition
1)Enzyme forms complex with substrate
2)Steady state level of complex does not change (assuming substrate concentration is in excess, i.e.: enzymes are saturated)
3)Only initial rates are measured so the back (reverse) reaction is negligible (not enough product has formed initialy). |
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Term
| Time Course of Enzyme-catalyzed Reaction |
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Definition
1)[Enzyme-substrate] over time increases and plateaus.
2)Free [enzyme] decreases and plateaus over time
3)Product increases over time.
4)E rapidly combines with S and a steady-state prevails in which the rate of S binding to E equals the rate of P leaving E.
5) The ratio of [ES]/[Efree] is proportional to [S] |
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Term
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Definition
vo=kcat[ES]
We cannot measure [ES]
Vmax occurs when enzyme is saturated so [ES]=[Etotal]
Vmax=kcat[Etotal] |
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Term
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Definition
vo = Vmax[S]/(kM + [S])
Vmax=kcat[Etotal]
kcat=Vmax/Etot (rearrangement; turnover number = S molecules converted to P molecules per enzyme active site per unit time.)
[S]=starting substrate concentration
Km, Michaelis constant. |
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Term
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Definition
Substrate concentration at which enzyme has half-maximal velocity.
Hyperbola curve. Km is NOT a derived dissociation constant, Kd (smaller = tighter binding) BUT the smaller value means it takes less substrate to bind 50% of the enzyme. |
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Term
| LB double reciprocal plot |
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Definition
1/vo = KM/Vmax(1/[S]) + (1/Vmax)
slope:KM/Vmax
y-intercept:1/Vmax
x-intercept:-1/Km |
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Term
| Km, Isozymes, Utilization of Glucose |
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Definition
2 Isozymes:
1) Hexokinase (efficient under normal conditions)
Catalyzes first step in glucose utilization (Glucose+ATP-->Glucose-6-phosphate+ADP), low Km for glucose, 0.05mM and tightly regulated (is inhibited allosterically by its product glucose-6-phosphate).
2) Glucokinase (hepatocytes) catalyzes same rxn (more efficient following a meal, storing energy)
Km of 10mM, much higher and is NOT inhibited by its reaction product. |
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Term
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Definition
Turnover number.
moles of product produced per second per mole per enzyme active site. Determined by measuring Vmax at known concentration of enzyme active sites. |
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Term
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Definition
Regulation of an enzyme can occur via a change in the
1)AMOUNT
2)LOCATION
3)ACTIVITY
of the enzyme
Changing ACTIVITY is faster than changing AMOUNT and is often seen in signal transduction and enzyme cascades. |
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Term
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Definition
You can change the intrinsic activity of enzyme by
1)Binding of Allosteric Effectors
2)Covalent modification
3)Binding of a “partner” protein
4)Proteolytic activation of an inactive precursor |
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Term
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Definition
1)Ligands that ONLY Binds to allosteric enzymes in sites other than the active site. Induces conformational change in enzyme so enzyme affinity for other ligand changes.
2)Can be positive or negative effectors.
3)Can be divided into
a)K Class (affects Km but not Vmax)
b)V Class (affects Vmax but not Km)
4)Most allosteric enzymes are oilgomeric (consists of multiple subunits)
5)Can either undergo
a)homotropic interactions (always +, increases Kcat)
b)heterotropic interactions (can be + and -) |
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Term
| Allosteric Enzyme Regulation (more detailed) |
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Definition
Enzymes do NOT follow MM kinetics
1)Sigmoid curve of homotropic enzyme. More efficient as [S] increases
2)K class, Vmax is not changed, + modulator=stepper vo curve and vice versa
3)Very rare, Vmax is altered, Km stays the same. |
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Term
| Aspartate Carbamoyltransferase (ATCase) |
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Definition
K-type Example.
1) Key enzyme in pyrmidine synthesis and sensitive to Pyr need
a)CTP inhibits enzyme (high CTP=high Pyr and stabilizes the T state)
b)ATP activates enzyme (high ATP means energy rich and high purines=need pyrmidines, binds to Reg. subunits and stabilizes the R state). |
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Term
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Definition
cAMP-dependent Protein Kinase
1)Regulatory subunits have pseudo-substrate region that binds to Catalytic (C) subunit in the absence of cAMP.
2)In the PRESENCE of cAMP, conformation of R subunits change so pseudo-substrate region can no longer bind, resulting in release of C subunits that are no active.
3)V-type allosteric enzyme. |
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Term
| Enzyme Regulation by Covalent Modification |
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Definition
Kinase phosphorylate proteins (which can activate or deactivate proteins).
Phosphatase dephosphorylate proteins.
AAs (hydrophilic aromatic AAs except for Ser)
Tyr
Ser* Most often.
Thr*
His |
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Term
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Definition
1)Pancreatic enzymes stored as inactive precursors (proenzymes) in zymogen granules before secretion into intestine through pancreatic duct.
2)Pepsin A is stored as proenzyme(Pepsinogen A) in the stomach, autoactivates itself at low pH.
3)Enzymes can activated by proteolysis (rxns occur within lumen of intestine)
4)Even though activation is irreversible, the enzyme's activity can be held in check by specific inhibitors. |
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Term
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Definition
1)Reversible
a)competitive (increases Km but has no effect on Vmax, can overcome with excess substrate) e..g: malonic acid is a competitive inhibitor of succinic acid.
b)noncompetitive (similar to negative effector allosteric enzyme except the focus is on inhibition). Do not resemble S, EIS complexes are also possible in addition to EI however EIS transforms S to P LESS efficiently and thus there is a decrease in Vmax.
c)tight binding
2)Irreversible |
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Term
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Definition
1) Resemble transition state and not S.
2) Ki is much lower than Km for the S
3) Reversible
4) Examples:
a)HIV Protease inhibitors
b)Statins-HMG-CoA reductase inhibitors
c)Methotrexate-Dihydrofolate reductase inhibitor |
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Term
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Definition
Bind covalently to enzyme and cannot be removed.
1)Flurouracil: Special type of irreversible inhibitor and cancer therapy drug.
2)Penicillin: Antibotics
3)Aspirin: inhibits cyclooxygenase (prostaglandin synthesis) |
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Term
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Definition
| "SUICIDE INHIBITOR". When Thymidylate Synthetase acts on it, enzyme is irreversibly inactivated. C-F bond is KEY and CANNOT be cleaved. Without Pyr, cells can't divide. |
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Term
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Definition
1)Transition State Analog.
Inhibits transpeptidase that crosslinks bacterial cell wall peptidoglycan strands, the last step in bacterial cell wall synthesis.
2)Strained 4-membered lactam ring that mimics the transition state of the normal substrate.
3)When Penicillin binds to active site, it forms a covalent bond with a Ser. |
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Term
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Definition
1)Inhibits cyclooxygenase activity by acetylating Ser 530 which blocks access of the S (arachidonate) to the active site of the enzyme.
2)Other nonsteroidal anti-inflammatory drugs NSAIDs, such as ibuprofen & indomethacin, inhibit cyclooxygenase activity by REVERSIBLY blocking the arachidonate binding site. |
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Term
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Definition
The TTP formed from the supplied uridine inhibits Carbamoyl-P synthetase by
FEEDBACK INHIBITION, resulting in a decrease in orotic acid production. |
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