ENZYMES. History of Enzymes. -1700s and early 1800s, the digestion of meat by stomach secretions and the conversion of starch to sugars by plant extracts and saliva were known. --mechanism by which this occurred had not been identified. History of Enzymes.
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-1700s and early 1800s, the digestion of meat by stomach secretionsand the conversion of starch to sugars by plant extracts and saliva were known.
--mechanism by which this occurred had not been identified.
-19th century, when studying the fermentation of sugar to alcohol by yeast, Louis Pasteur came to the conclusion that it was catalyzed by a vital force contained within the yeast cells called "ferments", which were thought to function only within living organisms.
--He wrote that "alcoholic fermentation is an act correlated with the life and organization of the yeast cells, not with the death or putrefaction of the cells.
Human pancreatic amylase
PROENZYME OR ZYMOGEN
Nitrogenase enzyme with Fe, Mo and ADP cofactors
(a) the substrate (xanthineoxidase)
(b) the source of the enzyme (pancreaticribonuclease)
(c) its regulation (hormone-sensitive lipase)
(d) a feature of its mechanism of action (cysteine protease)
Based on catalyzed reactions, the nomenclature committee of the International Union of Biochemistry and Molecular Biology (IUBMB) recommended the following classification:
Three major regulatory chemical reactions. (a)Acetylation - addition of an acetyl group to lysine's R group by acetyltransferase. (b)Methylation - addition of a methyl group to DNA's base (e.g. cytosine) by methylase. (c)Phosphorylation - addition of a phosphate group to the R group of tyrosine, serine or threonine (only tyrosine is shown here) by protein kinase.
3. HYDROLASES Catalyze hydrolysis reactions where a molecule is split into two or more smaller molecules by the addition of water
The role of rotamase and protein disulfide isomerase (PDI). The reactions catalyzed by the two enzymes can assist a peptide chain to fold into a correct three-dimensional structure
The IUBMB committee also defines subclasses and sub-subclasses
The molecule acted upon
a unique geometric shape that is complementary to the geometric shape of a substrate molecule
Lock and Key Theory
Enzyme may be used again
Postulated by Daniel Koshland
It states that, when substrates approach and bind to an enzyme they induce a conformational change
This change is analogous to placing a hand (substrate) into a glove (enzyme)
Hexokinase (a) without (b) with glucose substrate
This is a representation of carboxypeptidase A with a substrate (turquoise) bound in the active site. The active site is in the induced conformation.
This is a molecular model of the unbound carboxypeptidase A enzyme
A. CATALYSIS BY PROXIMITY
a) bimolecular reaction(high activation energy, low rate)
The field of biochemistry concerned with the quantitative measurement of the rates of enzyme-catalyzed reactions and the systematic study of factors that affect these rates
whereS is the substrate
E is the enzyme
ES is the enzyme-substrate complex
k1, k-1, and k2 are rate constants
Km + S
whereVo = initial reaction velocity
Vmax = maximal velocity
Km = Michaelis constant (k-1 + k2)/k1
S= substrate concentration
1. Relative concentrations
of E and S
2. Steady-state assumption
3. Initial velocity
a. Small Km
b. Large Km
Small Km for enzyme 1 reflects a high affinity of enzyme for the substrate
Large Km for enzyme 2 reflects low affinity of enzyme for the substrate
2. Relationship of velocity to enzyme concentration
3. Order of reaction
At high concentration of substrate([S]>>Km), The velocity of the reaction is zero order – that is, constant and independent OF substrate concentration
At low concentration of substrate([S]<<Km), The velocity of the reaction is first order – that is, proportional to substrate concentration
Also called a double-reciprocal plot
If 1/v0 is plotted VS 1/[S], a straight line is obtained
The intercept on the x-axis is equal to -1/Km
The intercept on the y-axis is equal to 1/Vmax
TYPES OF INHIBITION:
Inhibitor binds reversibly to the same site that the substrate would normally occupy, and therefore competes with the substrate for that site
Inhibitors tend to resemble the structures of a substrate, and thus are termed as substrate analogs
Malonate (¯OCOCH2COO¯) competes with Succinate (¯OOCCH2CH2COO¯) for the active site of succinate dehydrogenase (SDH)
SDH catalyze the removal of one H atom from each of the 2 methylene C’s of succinate
Malonate – Enzyme Complex
Vmax is unchanged: At high levels of substrate all of the inhibitor is displaced by substrate.
Km is increased: Higher substrate concentrations are required to reach the maximal velocity.
Inhibitor and substrate bind at different sites on the enzyme
The inhibitor binds to both E and ES
The noncompetitive inhibitor binds to an allosteric site (different location than the active site) of an enzyme
The binding of an inhibitor to the allosteric site alters the shape of the enzyme, resulting in a distorted active site that does not function properly.
Vmax is decreased: At high levels of substrate the inhibitor is still bound.
Km is not changed: Noncompetitive inhibitors do not interfere the binding of substrate to enzyme
I. SUBSTRATE CONCENTRATION
The rate of enzyme-catalysed reactions increases as the temperature rises to theoptimum temperature
Above a certain temperature, activity begins to decline because the enzyme begins to denature
Enzymes are usually damaged
above about 45°C
Each enzyme has an optimal pH
In order to interact, the E and S have specific chemical groups in ionized or unionized state
Amino group in protonated form (-NH3+) increase catalytic activity
At alkaline pH, amino group is deprotonated decrease in rate of reaction
Extremes of pH can lead to denaturation
REGULATION OF ENZYME ACTIVITY
A. ALLOSTERIC REGULATION
B. REGULATION OF ENZYMES BY COVALENT MODIFICATION
Enzyme Activity is Often Regulated
Feedback inhibition - a common form of enzyme regulation in which the product inhibits the enzyme .
ENZYMES IN CLINICAL USE
Enzyme inhibitors as DRUGS
Enzymes in CLINICAL DIAGNOSIS
2 GROUPS OF PLASMA ENZYMES
Elevated enzyme activity in the plasma may indicate tissue damage accompanied by increased release of intracellular enzymes, thus useful as a diagnostic tool
Elevated levels of ALT (alanine aminotransferase; also called glutamate: pyruvate transaminase; GPT) signals damage