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Regulation of Enzyme Systems

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systems are controlled by. regulating some of their enzymes. Types of Regulation ... processed into active enzymes; pieces of. polypeptide removed- IRREVERSIBLE ... – PowerPoint PPT presentation

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Title: Regulation of Enzyme Systems


1
Regulation of Enzyme Systems ?Metabolic systems
are controlled by regulating some of their
enzymes. ?Types of Regulation 1. Control of
gene expression gene? protein- SLOW,
IRREVERSIBLE 2. Zymogen activation inactive
proteins processed into active enzymes pieces
of polypeptide removed- IRREVERSIBLE 3.
Regulatory enzymes- complex enzymes REVERSIBLE
2
Zymogen Activation ?inactive precursor(zymogen) ?
ACTIVE - at cellular level ?
zymogen granules (full of zymogen proteins)
At the molecular level Pepsinogen
pepsin Trypsinogen trypsin Chymotrypsinogen
chymotrypsin
3
Details of Chymotrypsinogen Activation
?zymogen chymotrypsinogen processed by the
pancreas with chymotrypsin secreted into the
intestines. ?Chymotrypsinogen is a single
polypeptide of 245 aa residues. ?The active
product is sometimes called a-chymotrypsin
4
Chymotrypsinogen SS
SS INACTIVE 1 15 146 245
trypsin
SS
SS p-chymotrypsin ACTIVE 1 15 16
146 245
p- or
a-chymotrypsin
SS SS a-chymotrypsin ACTIVE 1
13 16 146 149 245
NOTE dipeptides ser14-arg15 thr147-asn148
removed
5
Regulatory Enzymes Feedback Control
Consider the conversion of thr to ile. E1
E2 E3 ………. thr ? B ? C ? D ???? ile
threonine deaminase
1st enzyme (E1)
Final product ile inhibits first enzyme in
pathway
6
Feedback Inhibition ?Last or near last product
of synthetic pathway inhibits the first enzyme in
pathway. ?efficient ?common ?regulatory
enzyme. Regulatory enzymes are complex
nearly always have quaternary structure.
7
Threonine deaminase is an example of
an allosteric regulatory enzyme
ile
Ile acts at site distant from the
substrate binding site, causing the enzyme to
undergo conformational change inhibiting the
enzyme
thr
8
Allosteric Enzymes 1. Complex,
multimeric 2. Several binding sites
Catalytic sites-- substrate Regulatory
sites -- inhibitory or activator -- sometimes
several sites of each type 3. Conformational
change 4. Not obey Michaelis-Menten kinetics
9
Generally, for an allosteric enzyme, rate(v) vs
S is sigmoidal instead of hyperbolic.
v
sigmoidal
S
?Simple enzyme hyperbolic at low S, activity
fairly high ?Complex, allosteric sigmoidal
activity low at low S, increases greatly in
midrange ?Efficiency is better, more responsive
to S
10
Example aspartate transcarbamoylase ATCase
O O
UTP H2N C-NH2 HN-C-NH2 TTP,
asp O asp
CTP OPO O
carbamoyl carbamoyl aspartate
phosphate ?Pyrimidines and purines are
needed for RNA and DNA synthesis
pyridine nucleotides
feedback
11
?CTP inhibits ATCase. Signal that more than
sufficient CTP has been produced. ?Purines are
synthesized by another metabolic pathway
involving other reactions. ?ATP activates ATCase.
Signal that there are more than sufficient
purines to encourage pyrimidine production.
12
Kinetic Properties of ATCase
?No jnhibitor or activator present ?sigmoidal, not
hyperbolic ?always implies cooperative, allosteri
c
v
asp
Example of Homotropic Effect
?response of v to S as example
above. sigmoidal (cooperative) response of an
enzyme to S
13
Heterotropic Effect influence of binding of an
inhibitor or activator on the binding
of substrate and its reaction
CTP inhibits ATCase lower activity for a given
asp
v
CTP
asp
CTP binding site different from substrate binding
site Regulatory Site
14
ATP activates ATCase less asp required
to achieve same rate
ATP
v
asp
ATP binding site is different from sites for asp,
carbamoyl phosphate, or CTP.
15
Allosteric Effect binding of effector at one
site influences the binding at another
site. NOTE asp, CTP, ATP not resemble
each other.
16
The Structure of ATCase
?12 subunits 6 catalytic 2 trimers AND 6
regulatory 3 dimers
?can be separated with 1 M urea (rather mild)
into 2 C3 Active, hyperbolic kinetic curve, no
response to ATP or CTP. 3 R2 Inactive, binds to
ATP, CTP, not to asp.
17
Aspartate Transcarbamoylase (ATCase)
Binding of CTP to sites on regulatory subunits
closes conformation, concealing substrate from
active site
CTP binding at site
18
Monod-Wyman-Changeux Model for Allosteric Enzymes
MWC model symmetric model
T tense R relaxed (active)
T
R
?Subunits in either T or R state ?All subunits in
protein must be in same state No Mixed
states ?Equilibrium between states ?T R states
bind S but R state binds better
19
S
S
S
S
S
OR
?Binding of S shifts the equilibrium to the R
state. ?binding of S to 1st 2nd site is
better. ?at high S, more R form exists
enzyme more active. ?at low S, more T form
exists enzyme has low activity.
20
4. Covalent Modification ?can control
enzyme activity ?most common is phosphorylation -
reversible
?example of phosphorylation the enzyme
phosphorylase a in glycogen utilization glycogen
polymer of glucose in muscle
liver. glycogen function store release
glucose.
21
Breakdown of glycogen (glucose)n Pi ?
(glucose)n-1 glucose-1-Pi
Energy source
Enzyme for removing glucose-1-phosphate is
phosphorylase a, activated by phosphorylation.
Another another another glu-1-Pi is removed
Part of cAMP cascade
(Pi inorganic phosphate)
22
The Cyclic AMP Cascade
epinephrine
Outside
receptor
AC
Inside cell
G- protein
ATP cAMP PPi
AC adenyl cyclase
protein kinase A active protein kinase
A R2C2 R2 2C tetramer
dimer 2 monomers inactive
active adds Pi to proteins
from ATP
23
protein kinase A (C monomer,
active) phosphorylase kinase phosphorylase
kinase phosphorylase b phosphorylase a
uses ATP to phosphorylate
active
uses ATP to phosphorylate
?Phosphoryated form a is active ?Phosphorylation
changes quaternary structure of phosphorylase
24
Bioenergetics ?Brief review of
some basic thermodynamics ?Changes in biological
systems involve both energy and entropy. ?Entropy
is the degree of randomness or disorder. ?The
important energy in biological systems is the
enthalpy or heat content.
25
?For a chemical or biochemical process to occur
requires that either enthalpy,?H, decreases or
entropy, ?S, Increases or both. ?Gibbs Free
Energy or FREE ENERGY combines the two in a
useful way. ?G ?H - T?S ?So a chemical
or biochemical reaction to occur spontaneously
requires that the change in free energy is
negative. ??G can be measured.
26
RULES 1. ?G negative,
spontaneous exergonic 2. ?G 0, equilibrium 3.
?G positive, endergonic the reaction will go in
the opposite direction unless energy is added to
the system. Note this says nothing about the
rate of the reaction it may be too slow to
observe even with a negative ?G.
27
For a chemical or biochemical system
at equilibrium, a characteristic Standard free
energy change ?G can be defined. A B ? C
D CD products
AB reactants
?G -RT ln Keq where R gas constant, 8.31
J/mole-K T absolute temperature in degrees
Kelvin (K C 273)

Keq
28
Equilibrium Approximate Standard Constant
Free Energy Change 0.001
17.1 kJ/mole 0.01 11.4
0.1 5.7 1 0
10 -5.7 100 -11.4 1000
-17.1
Question which direction will each go?
29
Tables of Standard Free Energies for All
Important Reactions Exist. ?Valuable information
about reactions
Standard Free Energy concept requires idea of
Standard Conditions.
1. pH 7 2. Ignore H2O
3. Starting Concentration is 1M for All
Compounds. So ?G is free energy starting at 1M
allowing rxn to go to equilibrium.
?Eqns exist to account for real concentrations
30
Free energies are additive so reactions can
be coupled.
?G (kJ/mole) 1) glucose Pi ?
glucose-6-Pi 13.8 2) ATP
? ADP Pi -30.5 glucose ATP
? glucose-6-Pi ADP -16.7 Note 1) is not
spontaneous 2) is very exergonic This coupling
reaction is carried out by enzyme hexokinase.
Hydrolysis of ATP is said to drive rxn. Driving
of rxn by ATP hydrolysis quite common. Coupling
A reaction with a negative free energy can drive
a reaction with a positive free energy.
31
High Energy Compounds ATP ?ATP is the
currency of chemical energy within cells. ?ATP is
the major chemical energy link between energy
yielding reactions such as the catabolism of
carbohydrates and fats, which generate ATP and
the energy requiring reactions, which utilize ATP.
32
Structure of ATP Note structures of ATP, ADP
AMP ? acid anhydride ?at pH 7.0 ATP ADP
are multicharged anions ATP-4 and ADP-3. ?Both
are nearly fully ionized.
?? ATP?? ADP?? AMP
ATP ADP are both found complexed to
Mg2, e.g., MgATP-2 and MgADP-.
33
Hydrolysis of ATP has large negative standard
free energy and is a high energy yielding
reaction. The hydrolysis is essentially
irreversible. So is the hydrolysis of ADP. AMP
hydrolysis not high energy ATP ? ADP Pi
?G -30.5 kJ/mole ADP ? AMP Pi
?G -30.5 kJ/mole AMP?Adenosine Pi ?G
-13.8 kJ/mole The free energy of hydrolysis in
the cell better because the concentration much
less than 1M.
34
Other compounds have high energy
yielding hydrolysis
phosphocreatine -- in muscle cells O H
CH3 O -OP-N-C-N-CH2-C-O- -O
NH2 ?G -43.1 kJ/mole Phosphoenolp
yruvate O O -O-P-O-C-C-O-
-O CH2
?G -62 kJ/mole
35
Bioenergetics is
important to predict and understand the structure
of metabolic pathways, the requirements for
energy harnessing coupling and the flux through a
pathway.
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