6 - THE BEHAVIOR OF PROTEINS: ENZYMES Flashcards
a biological catalyst
Enzyme
with the exception of some RNA’s that catalyze their own splicing, all enzymes are?
Proteins
can increase the rate of a reaction by a factor of up to 10^20 over an uncatalyzed reaction
Enzymes
some enzymes are so specific that they catalyze the reaction of only one stereoisomer; others catalyze a family of similar reactions
rate of reaction depends on its activation energy, DG
an enzyme provides an alternative pathway with a lower activation energy
most important function of proteins; performed by the protein, enzyme
Catalysis
highly specific; can distinguish isomers of a given compound
Enzymes
Biochemical reaction rates increase with temperature up to a point, but are limited due to enzyme denaturation, thermal inactivation of biomolecules, and the need to operate within an optimal temperature range for living organisms. Beyond these limits, reaction rates decrease as enzymes lose their functionality and other biological structures become compromised.
What are the two models that were developed to describe the formation of the enzyme-substrate complex?
Lock-and-key model
Induced fit model
substrate binds to that portion of the enzyme with a complementary shape; shape of the substrate and the conformation of the active site are complementary to one another
Lock-and-key model
binding of the substrate induces a change in the conformation of the enzymes that results to a complementary fit
Induced fit model
enzyme: undergoes a conformational change upon binding to the substrate
shape: active site becomes complementary to the shape of the shape substrate only after the substrate binds to the enzyme
Induced-fit model
What happens after the enzyme-transition complex is formed?
Catalysis occurs
In the complex:
S: bound close to atoms and is placed in the correct orientation with respect to the atoms with which it is to react; both proximity and orientation speed up the reaction
As some bonds are broken and new ones are formed, S —> product
Enzyme then S —> product
study on the formation of product from substrate
- catalyzes preferential hydrolysis of peptide bonds at residues containing aromatic side chains
- cleaves at other sites (lys, his, and gln) at relatively lower frequency
- catalyzes hydrolysis of ester bonds
Chymotrypsin
during hydrolysis of p-nitrophenyl ester by chymotrypsin,
- rate is dependent on the substrate
- at low [S], rxn rate increases with increasing [S]
- at higher [S], rxn rate changes very little, and a max. rate is reached
- graph: hyperbolic
carbamoyl phosphate + aspartate –> carbamoyl aspartate + hydrogen phosphate
- the 1st step in he pathway leading to the formation of CTP and UTP, which are ultimately needed for biosynthesis of RNA and DNA
- rxn rate also depends on [S] which is asp; graph is sigmoidal
Aspartate transcarbamoylase (ATCase)
- an allosteric protein
- similar to the kinetic behavior of hemoglobin
- allosteric proteins -proteins in which subtle changes at one site affect structure and function at another site
ATCase
non-allosteric proteins
Chymotrypsin and myoglobin
- 1913, Lenor Michealis and Maud Menten
- basic model for nonallosteric enzymes
Michealis-Menten
Enzymatic conversion of a substrate to a product,
S —> P
When the rate of the rxn is half its max. value, the substrate conc is equal to the Michealis constant
a substance that interferes with the action of an enzyme and slows the rate of a reaction; decreases the rate of an enzyme-catalyzed reaction
Inhibitor
a substance that binds to an enzyme to inhibit it but can be released
Reversible inhibitor
What are the two reverse inhibitors?
Competitive and noncompetitive
binds to the active (catalytic) site and blocks access to the substrate
Competitive inhibitor
compete with the substrate for binding to the enzyme’s active site; block the S access to it, thus competing with S for the E active site
Competitive inhibitor
bind to a site other than the active site, causing a change in the enzymes in the enzyme’s shape and function
cannot be overcome by increasing substrate conc
Noncompetitive inhibitor
binds to a site other than the active site; inhibits the enzyme by changing its conformation
Noncompetitive inhibitor
a substance that causes inhibition that cannot be reversed; usually involves the formation/breaking of covalent bonds to or on the enzyme
Irreversible inhibitor
similar to noncompetitive, but binding of inhibitor affects binding of S and vice versa
Mixed competitive
bind only to the enzyme-substrate complex, preventing the complex from releasing products
Uncompetitive inhibitors
What are the six major classes of enzymes?
- Oxidoreductases (EC 1)
- Transferases (EC 2)
- Hydrolases (EC 3)
- Lyases (EC 4)
- Isomerases (EC 5)
- Ligases (EC 6)
systematically governs enzymes classification and nomenclature
Enzyme commission (EC)
- catalyze oxidation-reduction reactions, where electrons are transferred from one molecule (the reductant) to the (the oxidant)
- e.g. alcohol dehydrogenase (EC 1.1.1.1)
Oxidoreductases (EC 1)
- transfer functional grps (e.g., a phosphate, methyl, or glycosyl grp) from one molecule to another
- e.g. hexokinase (EC 2.7.1.1)
Transferases
- catalyze the hydrolysis of various bonds, breaking molecules down using water
- e.g. lipase (EC 3.1.1.3)
Hydrolases
- catalyze the addition/removal of grps to form double bonds or break double bonds w/o hydrolysis or oxidation
- e.g. pyruvate decarboxylase (EC 4.1.1.1)
Lyases (EC 4)
- catalyze the rearrangement of atoms within a molecule, facilitating structural changes (isomerization)
- e.g. phosphoglucose isomerase (EC 5.3.1.9)
Isomerases (EC 5)
- catalyze the joining of two molecules with the formation of new chemical bonds, usually accompanied by the hydrolysis of ATP
- e.g. DNA ligase (EC 6.5.1.1)
Ligases (EC 6)
EC X.X.X.X
X = class, subclass, sub-sub-class, specific enzyme
refers to an enzyme that breaks down sucrose
Sucrase
The binding of oxygen by myoglobin is an example of a [] and is a []
non-allosteric behavior; non-allosteric protein
the binding of oxygen by hemoglobin is an example of [] and is an []
allosteric behavior; allosteric protein
What are the examples of allosteric proteins?
Their behaviors exhibit cooperative effects caused by subtle changes in quaternary structure
ATCase and hemoglobin
catalyzes the 1st step in a series of reactions in which the end product is cytidine triphosphate (CTP), a nucleotide triphosphate needed to make RNA and DNA
ATCase
are energetically costly and involve many steps
pathways that produce nucleotides
an excellent example of how such a pathway is controlled to avoid the overproduction of such compounds
The reaction catalyzed by aspartate transcarbamoylase
an efficient control mechanism because the entire series of reactions can be shut down when an excess of the final product exists, thus preventing the accumulaiton of intermediates in the pathway
Feedback inhibition
study on the reaction catalyzed by ATCase leads eventually to the production of CTP
an inhibitor of ATCase; an example of feedback inhibition (end-product inhibition), in which the end product of the sequence of reactions inhibits the first reaction in the series
CTP
an oligomer whose biological activity is affected by other substances binding to it; change the enzyme’s activity by altering the conformation(s) of its quaternary structure
allosteric enzyme
a substance that modifies the behavior of an allosteric enzyme
allosteric effector
an allosteric effector can be?
allosteric inhibitor
allosteric activator
Two principal models for the behavior of allosteric enzymes?
Concerted model and sequential model
The downstream products combined to the regulatory site causing the enzyme to change shape and prevent it from binding to its substrate; causes the entire pathway to halt and new products are no longer formed
Conversely, when the amount of downstream product gets too low, the enzymes interact with their usual substrates and begin forming their usual products again
Feedback inhibition plays a critical role in preventing cells from wasting resources and energy
what does the sigmoidal shape of the curve describes?
Allosteric behavior
The binding site for ATP on the enzyme molecule is the same as that for CTP, but ATP is rather an INHIBITOR like CTP. When CTP is in short supply in an organism, the ATCase reaction is not inhibited, and the binding of ATP increases the activity of the enzyme still more.
proposed in 1965; has the advantage of comparative simplicity
Concerted model
The enzymes has two conformations:
R (relaxed) - binds substrate tightly; active form
T (tight or taut) - binds the substrate less tightly; inactive form
Concerted model
proposed in 1966; gives a realistic picture of the structure and behavior of proteins
Sequential model
change in conformation is induced by the fit of the substrate to the enzyme, as per the induced-fit model of substrate binding
represents cooperativity
Sequential model
binding of the substrate induces a conformational change from the T form to the R form
Sequential model
most enzyme molecules are in the T (inactive) form in the?
Absence of substrate
What shifts the equilibrium from the T (inactive) form to the R (active) form
The presence of substrate
In changing from T to R and vice versa, all subunits change conformation simultaneously; all changes are concerted
In the concerted model for allosteric behavior,
- the binding of substrate, inhibitor, or activator to one unit shifts equil. b/w R form of the enzyme (binds substrate strongly), and T form (does not bind substrate strongly)
- conformational change takes place in all subunits at the same time
In the sequential model,
- the binding of the substrate induces the conformational change in one subunit, and the change is subsequently passed along to other subunits
is one whose activity is modulated by molecules binding at sites other than the active site
Allosteric enzymes
What is the allosteric enzyme of ATCase?
CTP (cytidine triphosphate)
end product of the pathway in which ATCase is involved, binds to the enzyme and inhibits its activity through feedback inhibition
CTP (cytidine triphosphate)
Why is ATCase an allosteric enzyme?
because it can be regulated by molecules like CTP, which binds at allosteric sites and inhibits its catalytic activity through a conformational change, thereby controlling the flow of metabolites in pyrimidine biosynthesis
Why is chymotrypsin not considered an allosteric enzyme?
Because it does not exhibit allosteric regulation, where the binding of a molecule at a site other than the active site affects the enzyme’s activity
inactive precursor of an enzyme where cleavage of one or more covalent bonds transforms it into the active enzyme
Zymogen
- synthesized and stored in the pancreas
- a single polypeptide chain of 245 amino acid residues cross-linked by five disulfide (-S-S-) bonds
- when secreted into the small intestine, the digestive enzyme trypsin cleaves a 15-unit polypeptide from the N-terminal end to give the pi-chymotrypsin; acts on itself to remove two dipeptide fragments, producing alpha-chymotrypsin (fully active)
Chymotrypsinogen
non-protein chemical compounds/metallic ions that help enzymes perform their catalytic functions
- inorganic ions (Mg2+ [for DNA polymerase], Fe2+, Zn2+ [for alcohol dehydrogenase]
- organic molecules (coenzymes are a subset of organic cofactors)
stabilize enzyme structure, participate in the actual catalytic reaction, or assist in substrate binding
Cofactors
specific type of organic cofactor; typically small organic molecule, that helps enzymes catalyze reactions by acting as carriers for chemical groups or electrons
organic molecules often derived from vitamins
often participate in the reaction by temporarily accepting or donating atoms, ions, or FG
usually regenerate after rxn, allowing them to be used repeatedly
Coenzyme
All coenzymes are cofactors, but not all cofactors are coenzymes (search more on this)
Coenzymes often participate directly in the enzymatic reaction, while some cofactors may play more structural or stabilizing roles.
