4 enzyme mechanism 20140906
DESCRIPTION
HKU science lecture notesTRANSCRIPT
Mechanisms of enzymes
Energy diagram for a single-step reaction (A-B + C → A + B-C)
(Transition state)
Activation energy
(required for reactants to achieve transition state)
Effect of an enzyme on a chemical reaction
- Lowering of activation energy - More reactants (= substrates) achieve transition state - Acceleration of reaction
•Enzyme binding of substrates increases the initial ground state of the enzyme-substrate complex (brings reactants in close proximity and into correct orientation, thus more transition state formation)
•Stabilize the transition state (by tight binding) to lower activation energy barrier.
How do enzymes reduce the activation energy?
Induced Fit - Substrate-induced conformation change - A substrate specificity effect - Example: Hexokinase
Open form: no substrates Closed form: with bound glucose
- Exclusion of water - No hydrolysis of ATP
- Not a simple “lock and key” model - Dynamic interaction between enzyme and
substrates
Chemical modes of enzymatic catalysis
Polar amino acid residues in active sites
- In the enzyme-substrate complex, substrates are in proximity to reactive amino acid resides in enzyme active sites
- Enzymes usually have 2-6 catalytic residues in active site - Ionizable side chains (in polar AA) can act as acids, bases, or nucleophiles - Involved in substrate binding and/or transition state formation
1. Acid-Base Catalysis
- A proton is transferred between the enzyme and the substrate - Amino acid side chains that can act as acid-base catalysts:
- Serving as proton donor or accepters depending on their protonation status
• Involves AA residues that can accept a proton
• Can remove proton from –OH, -NH, -CH, etc. and cause bond cleavage
• Creates a strong nucleophillic reactant (i.e. X:-)
Base catalysis:
• Removes a proton from water
• Generates an OH- equivalent which attacks the carbonyl carbon
• Cleavage of C-N bond
1.
2.
Acid catalysis
- Proton donation - A covalent bond may break more easily if one of its atom is protonated
BH+
2. Covalent Catalysis (nucleophilic catalysis) • 20% of all enzymes employ covalent catalysis
A-X + B + E BX + E + A
• A group from a substrate binds covalently to enzyme
• The intermediate enzyme substrate complex (A-X) then donates the group (X) to a second substrate (B)
• Side chains that can act as covalent catalysts:
They serve as nucleophiles in the deprotonated forms
Effect of pH on enzyme reaction rates
- Ionizable side chains of catalytic AA residues require proper pH for achieving the necessary protonated status for catalysis
- Catalytic cysteine-25 and histidine-159 residues in papain (a papaya protease):
The activity of papain depends on His-159 and Cys-25 in the active site
General acid-base catalysis mechanism for triose phosphate isomerase
Formation of enzyme-bound intermediate:
Glu-165: a general acid-base catalyst His-95: shuttles a protein between the oxygen atoms of an enzyme-bound intermediate
- cleavage of peptide bond - a serine protease - Catalytic triad: His-57, Asp-102, Ser-195
Acid-base catalysis and covalent catalysis for chymotrypsin
Chymotrypsin
Peptide substrate
scissile bond
specificity pocket
Structural similar serine proteases with different substrate specificities
Chymotrypsin Trypsin Elastase
Specificity pockets of three serine proteases:
C
O
Cα
(attacked by Ser-195)
N