Enzyme Regulation, Allostery and Inhibition

Do enzymes alter the reaction rate or the reaction equilibrium

reaction rate

regulated enzyme

enzyme with catalytic activity that increases or decreases in response to changing conditions in the cell or organism

substrate

Many enzymes have a Km that is near to the concentration of their what that
is found in cells

regulated enzymes are modulated in a variety of ways:
– reversible what modification

covalent

proteins

regulated enzymes are modulated in a variety of ways:
– binding of separate regulatory what

cleavage

regulated enzymes are modulated in a variety of ways:
– removal of peptide segments by proteolytic what

allostery

regulation through reversible, noncovalent binding of small compounds called allosteric modulators or allosteric effectors

phosphorylation

one of the most common types of covalent (but reversible) modification of proteins
and enzymes is what

protein kinases

catalyze the attachment of phosphoryl groups to specific amino acid residues

protein kinases catalyze the attachment of phosphoryl groups to specific amino acid residues
- which in eukaryotes

Ser and Thr, or Tyr

protein kinases catalyze the attachment of phosphoryl groups to specific amino acid residues
- which in prokaryotes

His

protein phosphatases

remove phosphoryl groups from proteins

phosphate

These are the three amino acids that can be modified by addition of a what group in eukaryotes, when proteins are ‘phosphorylated

adrenaline

Much of our knowledge of signal transduction came from 20th C studies on the role of what on glycogen breakdown during the ‘fight or flight’ response

glycogen phosphorylase

the enzyme that cleaves glucose from the glycogen chain in the form of glucose-1-phosphate

phosphorylation

The activity of glycogen phosphorylase is controlled by what

glycogen phosphorylase is what when phosphorylase kinase puts a phosphate on it

activated

Conformational

Phosphorylation in Glycogen Phosphorylase Causes a what Change

active

Phosphorylation in Glycogen Phosphorylase Causes a Conformational Change, conformational change shifts enzyme into a more catalytically what state

Sites

Some Enzymes are Phosphorylated at Multiple what

phosphorylase kinase becomes active when it is phosphorylated by protein kinase what

A

subunit

protein kinase A becomes active when it is released from its regulatory what

Dissociation

Regulation of Enzyme Activity by what of a Regulatory Subunit

protein kinase A

? catalytic subunits (constitutively active)
? regulatory subunits (keep catalytical subunit off)

2,2

target

cAMP binding to the regulatory subunits causes the what to change conformational and release the catalytic subunits which are free to act on what proteins

zymogen

inactive precursor that is cleaved to form an active protease enzyme

proprotein/proenzyme

precursors that are cleaved to form other proteins

trypsin

for the protease chymotrypsin to be activated it is first cleaved by the protease what

itself

for the protease chymotrypsin to be activated it is first cleaved by the protease trypsin, then chymotrypsin is further processed by who

cascade

a process where the same thing happens several times in a row

amplification

cascade can allow ‘what’ – a large response to a small initial signal

Blood

A Cascade of Proteolytically Activated Zymogens Leads to what Coagulation

collagen

Blood clots occur when what is exposed to blood

fibrin

Blood clot are an aggregation of specialized cell fragments called platelets that is cross-
linked and stabilized by fibers consisting of the protein what

fibrinogen

Fibrin is derived from the soluble zymogen what

Activation

Two Regulatory Cascades Lead to Fibrinogen what

proteolysis

The pathways converge on the step in which inactive prothrombin is converted t oactive thrombin by what

serine

The active what protease thrombin then cleaves fibrinogen into fibrin

Allosteric Enzymes Are Usually Regulated by what of the Pathway They Act in or Control

Products

regulated

The activity of many pathways is what to meet the needs of the cell

However, instead of regulating the activity of all the enzymes, it makes more sense to regulate the activity of which enzyme in the pathway, or the enzyme that controls which committed/irreversible step in pathway

first

In this pathway, all the reactions are what, so once A → B, the final product F will be made.

irreversible

committed

In this pathway, all the reactions are irreversible, so once A → B, the final product F will be made. Therefore, the first step is the what step in the pathway

feedback

In this example, the amount of product F is regulated by what inhibition - the final product F feeds back to inhibit the first enzyme in the pathway

regulatory

The pathway product F inhibits enzyme e1, which controls the committed step in the pathway, by binding to a what site on the enzyme that is distinct from the active site

allosteric

The pathway product F inhibits enzyme e1, which controls the committed step in the pathway, by binding to a regulatory site on the enzyme that is distinct from the active site
This site is called the what site

allosteric

Enzyme e1 is therefore an what enzyme or an enzyme that shows what regulation

committed

allosteric enzymes’ often catalyze which step of pathways

steps

‘Michaelis–Menten’ enzymes facilitate the remaining what

Allosteric enzymes may be inhibited, stimulated, or both, by what, regulatory molecules that convey the metabolic needs of the cell

small

behavior

Two models, the concerted model and the sequential model, explain the what of allosteric enzymes

proteins

In these models (the concerted model and the sequential model), binding of substrate or inhibitor to one protein, alters the conformation of other what in the complex

Features of which model for allosteric regulation:
– the enzyme exists in two different structures, T (tense) and R (relaxed)

concerted

Features of the concerted model for allosteric regulation:
– the enzyme exists in two different structures, T (tense) and R (relaxed)
– the R state is more or less enzymatically active than the T state

more

same

Features of the concerted model for allosteric regulation:

all active sites must be in which state

the binding of substrate to one active site shifts all the other active sites into which state (a concerted conformational change

R

The sequential model is basically the same as the concerted model but proposes that subunits undergo sequential changes in conformation to the active which state (not all at once

R

Molecules

Regulator what Alter the T ⇌ R Equilibrium

Inhibitors stabilize which state

T

Activators stabilize which state

R

Allosteric regulation by substrates

homotropic

Allosteric regulation by substances that are not substrates

heterotropic

regulation in which the enzyme modulator is a small molecule that is not the substrate

heterotropic

modulator

Binding of the allosteric what (M) to the allosteric site shifts the enzyme into an active conformation (R) more likely to bind substrate or carry out catalysis

substrate

The reaction velocity of allosteric enzymes displays a sigmoidal (s shape) relationship to
what concentration

hyperbolic

The reaction velocity of allosteric enzymes displays a sigmoidal (s shape) relationship to
substrate concentration compared to Michaelis-Menten enzymes which are more what in their kinetic

An allosteric activator (+) may cause the velocity vs substrate curve to shift where and become more hyperbolic like an enzyme following Michaelis-Menten kinetic

left

An allosteric inhibitor (-) may cause the velocity vs substrate curve to shift to the what and become more flat

right

With allosteric inhibitors and activators, which is altered

Km

modulators

Allosteric what of enzyme activity allow the activity of pathways to be fine-tuned to the needs of the cell

aspartate transcarbamoylase (ATCase)

catalyzes the formation of carbamoylaspartate, an early step in pyrimidine biosynthesis

The nucleotide CTP is a what allosteric regulator of ATCase

negative

The nucleotide ATP is a what allosteric regulator of ATCase

positive

inhibited

Enzymes can be what so their catalytic activity is reduced or abolished

permanent

Inhibition can be reversible or what

inhibitors

~50% of prescribed drugs are what

Three common types of reversible inhibition

The inhibitor is structurally similar to the substrate and binds to the active site of the enzyme, preventing the actual substrate from binding

Competitive inhibition

Three common types of reversible inhibition
The inhibitor binds to only the enzyme–substrate complex (not to the free enzyme

Uncompetitive inhibition

Three common types of reversible inhibition

The inhibitor binds to a site different from the active site (does not compete with substrate) on either the enzyme (this is negative allostery) or enzyme–substrate complex

Noncompetitive inhibition

Inhibitor binds in active site

Competitive

Inhibitor does not bind in active site

Uncompetitive & Noncompetitive

inhibitor looks like substrate and binds enzyme at active site and competes with the normal substrate for binding

competitive

In presence of the inhibitor, Vmax of the enzyme is unchanged because the inhibition
can be overcome by doing what to the substrate (the substrate can ‘outcompete’

adding more

The KM is what in the presence of an inhibitor because it takes more substrate
to reach ½ VMAX

increased

As more competitive inhibitor is added, the rate of the reaction does what

decreases

But at each concentration of competitive inhibitor, if what substrate is added, the reaction approaches uninhibited VMA

more

As more competitive inhibitor is added, the KM what

increases

With competitive inhibitors does Vmax change

No

inhibitor binds the enzyme-substrate complex only

uncompetitive

The enzyme–substrate–inhibitor (ESI) complex does or does not form product

does not

uncompetitive

Consequently, Vmax is lower in the presence of inhibitor (you can’t add enough substrate to overcome which inhibition)

The KM is also what in uncompetive inhibition (for complicated reasons)

lower

As more uncompetitive inhibitor is added, the rate of the reaction does what

decreases

inhibitor can bind to free enzyme or to the enzyme–substrate complex, but not at the active site

noncompetitive

Vmax is what in the presence of a noncompetitive inhibitor. Noncompetitive
inhibition cannot be overcome by increasing substrate concentration (because the
substrate and inhibitor bind different sites

lower

bind

KM is not changed by the presence of a noncompetitive inhibitor because the substrate can still what its site.

As more noncompetitive inhibitor is added, the rate of the reaction does what

decreases

if more substrate is added, the reaction never approaches uninhibited VMAX.
VMAX is lower

uncompetitive

if more substrate is added, the reaction never approaches uninhibited VMAX

noncompetitive

Which inhibitors bind very tightly to enzymes and permanently reduce or abolish enzyme activity

Irreversible