Chemistry 501 Handout 6 Enzymes Chapter 6

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Chemistry 501 Handout 6EnzymesChapter 6
Lehninger. Principles of Biochemistry. by Nelson
and Cox, 5th Edition W.H. Freeman and Company
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With exception of a small group of catalytic RNA
molecules, all enzymes are proteins Largest class
of proteins. More than 3000 enzymes known.
Enzymes are biological catalysts that accelerate
reactions. Enzymes are generally highly specific
and react with only one substrate to form one
product and can enhance reaction rates by as much
as 1017.
Some enzymes require cofactors / coenzymes for
activity
Coenzymes act as transient carriers of specific
functional groups
A cofactor or coenzymes that is very tightly or
even covalently bound to the enzyme protein is
called a prosthetic group.
Complete, catalytically active enzyme
holoenzyme Only protein part (without cofactor
and/or coenzyme) apoenzyme or apoprotein
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Enzymes are classified by the reactions they
catalyze
International Classification System
(nomenclature)
(Four-part classification number and a systematic
name)

• Systematic name ATPglucose phosphotransferase
• Classification number 2.7.1.1.
• 2. --gt Transferase (class)
• 7. --gt phosphotransferase (subclass)
• 1. --gt phosphotransferase with a hydroxyl group
  as acceptor
• 1. --gt D-glucose as the phosphoryl group acceptor

ATP D-glucose --gt ADP D-glucose-6-phospate
Hexokinase
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Binding of a substrate to an enzyme at the active
site
Representation of a simple enzymatic reaction
Enzymes affect reaction rates, not equilibria
E S ? ES ? EP ? E P
Reaction coordinate diagram for a chemical
reaction
Reaction coordinate diagram comparing
enzyme-catalyzed and uncatalyzed reactions
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Reaction S ? P Reaction rate V k S
If the rate depends only on the concentration of
S (first-order)
From the Transition-State Theory
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Weak interactions between enzyme and substrate
are optimized in the transition state
The energy derived from enzyme-substrate
interaction is called binding energy, DGB
Enzyme active sites are complementary not to the
substrate per se, but to the transition state
through which substrates pass as they are
converted into products during an enzymatic
reaction
Complementary shapes of a substrate and its
binding site on the enzyme
Role of binding energy in catalysis
An imaginary enzyme (sticase) designed to
catalyze breakage of a metal stick
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How a catalyst circumvents unfavorable charge
development during cleavage of a amide
Rate enhancements by entropy reduction
Specific acid-base catalysis
General acid-base catalysis
Hydrolysis of an amide bond - same reaction as
that catalyzed by chymotrypsin and other
proteases
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Amino acids in general acid-base catalysis
Covalent and general acid-base catalysis
First step of the reaction
His57
His57
Amino acid side chains and the functional groups
of some cofactors can serve as nucleophiles in
the formation of covalent bonds with substrates
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Enzyme Kinetics
Effect of substrate concentration on the initial
velocity of an enzyme-catalyzed reaction
Michaelis-Menten kinetics
initial rate measurements
S gtgt E ES constant V0 K2 ES
Michaelis constant
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Dependence of initial velocity on substrate
concentration
When km gt gt S
When S gt gt km
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A double-reciprocal or Lineweaver-Burk plot
Lineweaver-Burk equation
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Kcat describes the limiting rate of any
enzyme-catalyzed reaction at saturation
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Many enzymes catalyze reactions with two or more
substrates
Common mechanisms for enzyme-catalyzed
bisubstrate reactions
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Pre-steady state kinetics can provide evidence
for specific reaction steps
Ternary complex is formed in the reaction (as
indicated by the intersecting lines)
Ping-Pong (double displacement) reaction
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Enzymes are subject to reversible and
irreversible inhibition
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Enzymes are subject to reversible and
irreversible inhibition
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Irreversible inhibition
Reaction of chymotrypsin with diisopropylfluoroph
osphate irreversibly inhibits the enzyme
Suicide inactivators or mechanism-based
inactivators
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Examples of Enzymatic Reactions
Chymotrypsin is a protease specific for peptide
bonds adjacent to aromatic amino acids (Trp,
Tyr, Phe)
Key active-site amino acid residues Ser195,
His57, and Asp102
Aromatic amino acid side chain
Pocket in which the amino acid side chain of the
substrate is bound
Three polypeptide chains linked by disulfide
bonds
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The chymotrypsin mechanism involves acylation
and deacylation of a Ser residue
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Regulatory Enzymes
Allosteric enzymes undergo conformational changes
in response to modulator binding
Conformational change
Subunit interactions in an allosteric enzyme, and
interactions with inhibitors and activators
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Two views of the regulatory enzyme aspartate
transcarbamoylase
This allosteric regulatory enzyme has two stacked
catalytic clusters, each with three catalytic
polypeptide chains (in shades of blue and purple)
and three regulatory clusters, each with two
regulatory peptide chains (in red and yellow).
Modulator binding produces large changes in
enzyme conformation and activity
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Feedback inhibition
Buildup of the end product ultimately slows the
entire pathway
In many pathways a regulatory step is catalyzed
by an allosteric enzyme
Example of heterotropic allosteric inhibition
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Phosphoryl groups affect the structure and
catalytic activity of enzymes
Regulation of muscle glycogen phosphorylase
activity by multiple mechanisms
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Some enzymes and other proteins are regulated by
proteolytic cleavage of a precursor
Activation of zymogens by proteolytic cleavage
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