Enzymes 1 and 2 Flashcards
Define simple enzyme, conjugated enzyme, apoenzyme, cofactor, coenzyme, holoenzyme, and proenzyme.
Simple: contains only the polypeptide portion.
conjugated: contains polypeptide portion and non-protein portion needed both in order to work.
apoenzyme: the protein portion of a CONJUGATED enzyme.
Cofactor: (prosthetic group) the non-protein portion of a CONJUGATED enzyme.
Coenzyme: organic factor. Ex. Vitamins. metals are also common cofactors
holoenzyme: the complete, native enzyme
proenzyme (zymogen): an inactive precursor of a native enzyme
Apoenzymes are NOT native enzymes b/c part of the polypeptide has to be removed to give a native enzyme
Know the major classes of enzymes and factors that effect enzymatic reactions.
Oxidoreductases = catalyze oxidation-reduction reactions (NADH)
Transferases = catalyze transfer of functional groups from one molecule to another.
Hydrolases = catalyze hydrolytic cleavage
Lyases = catalyze removal of a group from or addition of a group to a double bond, or other cleavages involving electron rearrangement.
Isomerases = catalyze INTRAmolecular rearrangement.
Ligases = catalyze reactions in which two molecules are joined.
What are the factors that effect enzymatic reactions?
Factors that effect the rate include: rate constants (k), concentration of reactant, and viscosity.* *
enzymatic rate depends on pH and Temparture.
Diffusion control: the overall rate of any process cannot exceed the probability of the maximum number of collisions b/w reaction components.
Understand the significance of Km and Vmax
Km: The Km is the concentration of substrate
at which 1/2 of the active sites are filled
As Km decreases, the power of the reaction increases
Vmax: it is the maximal rate of the reaction–but can NEVER be reached.
The speed of a reaction will increase
until the maximum processing rate of
the enzyme is reached = all the
active sites are filled and chugging away
To increase the speed of the reaction:
*add more enzyme
*improve k2
activator
engineer better active site
*If [S] is very large compared to Km then Vo=Vmax
If Km=[S] then Vo=Vmax/2
Understand irreversible, competitive and non competitive inhibitors and their importance in drug design.
Competitive Inhibition
Competitive inhibitor- inhibitor mimics substrate and attaches to same active site. reversible
ex. methanol and ethanol competing for active site on alcohol dehydrogenase
competitive inhibitor- inhibitor competes for active site. At higher levels of [S] inhibitor is “swamped out”
Vmax does NOT change, but Km INCREASES!! This is shown with a hyperbolic plot
-can form ES or EI complex
NON-COMPETITIVE INHIBITION
Non-competitive inhibitor- does NOT compete for same active site, but changes the active sites conformation when bonded. Large amount of substrate CANNOT overcome inhibitory effect
Non-competitive inhibition- as the number of inhibtors increases, the efficiency of processing the product decreases. higher [S] cannot “swamp out” effect. The problem is not at the active site. Km does NOT change and Vmax stays the DECREASES (the enzyme is less efficient at processing the product).
-can form ES, EI or ESI complex
**for both refer to the Line-Weaver Burke plot: 1/Vmax is the y-intercept and Km/Vmax is the slope of the line. The higher the Vmax the lower the y-intercept. highest line most inhibited
Importance of allosteric enzymes.
Catalyze irreversible reactions; are rate limiting
Generally contain more than one polypeptide chain (Quaternary Structure)
Do not follow Michaelis-Menten Kinetics
Can be upregulated by allosteric activators at constant [S]
Can be down regulated by allosteric inhibitors at constant [S]
Activators and Inhibitors need not have any structural resemblance to substrate structure
Irreversible inhibitors generally result in the destruction or modification of an essential amino acid required for enzyme activity. Frequently, this is due to some type of covalent link between enzyme and inhibitor. These inhibitors are designed to mimic the natural substrate in recognition and binding to an enzyme active site. Upon binding and some catalytic modification, a highly reactive inhibitor product is formed that binds irreversibly and inactivates the enzyme. Use of suicide inhibitors have proven to be very clinically effective
Major controls of enzyme activity
Major controls of enzyme activity
The availability of substrates and cofactors usually determines how fast the reaction goes. As product accumulates, the apparent rate of the enzymatic reaction will decrease due to lack of substrate.
Enzyme activity can be regulated through covalent modification.
Enzyme activity can be regulated allosterically.
Zymogens, isozymes, and modulator proteins may play a role.
Genetic regulation of enzyme synthesis and decay determines the amount of enzyme present at any moment.
Enzyme level: The enzyme may be activated or inhibited by either noncovalent or covalent interactions. This is the most rapid control system.
- Hormonal level: A hormone is secreted and carries a message to the cell and in turn an enzyme is activated or inhibited. The speed of this control system is intermediate.
- Gene level: A message is sent to the nucleus either express or repress a gene. This determines the amount of enzyme produced and is the slowest control measure.
Mechanisms of enzymatic reactions.
- enzymatic activity:
E+S= {ES} = E + P
- competitive inhibitor
(reversible)
E+ S= ES = E + P or
E+ I = EI= no rxn
- Non-competitive inhibitor
(reversible)
E+S= ES= E +P or
E+I= EI= no rxn
ES + I= ESI= no rxn
- Allosterics= irreversible!
Understand the role of the active site in enzymatic reactions.
Active Site = The region of the enzyme that binds and acts upon the substrate
binding = few or many weak attractions that can be reversed
ion ion, H-bond, van der Waals
any type of amino acid side chain can be used in binding but must have a molecular match with substrate
amino acids can be from anywhere in chain
acting upon = formation of a transient covalent intermediate that rearranges to product(s) = enzyme-substrate complex
coenzymes and cofactors are sometimes needed to effect catalysis
shape of active site = generally crevice or cavern on surface of enzyme
specificity of active site
low = broad = floppy site
high = narrow = tight site
models of binding at active site
Lock-and-Key Model
Active site has a rigid shape that is complementary to that of the substrate
Induced-fit Model
The shape of the enzyme adjusts to fit the proper substrate upon binding of the substrate
Absolute specificity
The enzyme catalyzes one reaction for one substrate.
Urease
Stereochemical Specificity
The enzyme catalyzes a reaction for only one stereoisomer.
L-amino acid oxidase
Group Specificity
The enzyme catalyzes a reaction involving only certain functional groups
carboxypeptidase
Linkage Specificity
The enzyme catalyzes a reaction for only a single type of bond.
phosphatases
