Download Serine Proteases

Serine Proteases
Oxyanion hole
O
Specificity
pocket
O
NH HN
NH2
H2N
H
N
H
O
N
H
substrate
O
HN
HO
O
HN
O
N
O
HN
O
Catalytic triad
NH
Components of the enzymatic pocket:
1. Catalytic triad
(asp-his-ser)
2. Oxyanion hole
3. Specificity pocket
Basic mechanism
O
N
NH HN
H2N
H
N
H
N
H H
NH2
O
N
H
substrate
H 2N
H
N
H
NH2
O
O
O
H O
H N
N
H2N
substrate
H
HN
NH
HN
O
O H
N
O
O
O
O H
O
O
H
HN
N H
O
N
H
N
O HO
O
HN
NH HN
O
O
NH2
O
O
O
N
OH HN
N
O
HN
HN
HN
O
O
O
O
O
O
O
NH HN
N
N
H H
NH2
H2N
H
N
H
O
NH2
OH
H2N
H
N
H
O
O
H O
H N
N
NH
HN
O
O
HN
O H
N
HN
N H
O
HN
O
OH
O
O
HN
O
O
O
Aspartyl Proteases
R
H
O
N
H
H
O
Asp
H
HO
O
Asp
substrate
Components of the enzymatic pocket:
1. Two aspartate residues
2. Enzyme bound water
3. Specificity for hydrophobic
amino acids
O
O
Charge repulsion of the two aspartate residues decreases side
chain acidity
Basic mechanism
-There is no enzyme-substrate intermediate!
R
O
H
N
H
H
O
R
substrate
H
O
H
N
HO H
Asp
O
R
H
O
OH
H2N
substrate
Asp
Asp
H
Asp
O
H O
O
substrate
O H
O
Asp
O
O
O
O
Asp
OH
O
• Be able to compare the mechanisms of the
two protease types:
Catalytic
residues
Catalytic triad
(asp-his-ser)
Dual aspartate
residues
Intermediate
Enyzme-bound
Tetrahedral,
geminal diol
Specificity
Basic amino
acids (arginine,
lysine
Nonpolar amino
acids (e.g.
valine, leucine)
Be able to use the mechanisms of these two
proteases to develop inhibitors
Serine proteases: make use of the enzyme bound
intermediate to covalently trap the enzyme
-identify inhibitors by basic amino acids and
good leaving groups/electrophilic sites
Aspartyl proteases: no enzyme bound intermediate
-mimic the tetrahedral nature of the bound
intermediate (transition state analog) NH
2
H
N
O
O
N
H
H
N
O
H
N
OH
N
H
OH
O
O
O O O
P
N
H
NH2
H2N
NH
O
H2N
N
H
H
N
O
NH
Ncyc
O
N
H
NH
H2N
O
H2N
NH2
H2N
H2N
O
O
Ncyc
N
H
NH2
H
N
O
NH
O
N
H
H
O
N-terminal hydrolases
• Examples include the proteasome, penicillin acylase, glutamine PPP
amidohydrolase
• Catalytic domain consists of a bound water molecule and an Nterminal serine, threonine, or cysteine
H+
O
H
O
N
H
H+
O
H O
H
B-
N
OH
H
O
H H2N
O
O
O
H
N
R
H
B-
O
H
N
OH
NH2
O
H
O
H O
O
HN
O
H2N
OH
O
R
O
H
H
N
H HN
H2
R
HN
R
R
O
H
NH2
Autoproteolysis
• Similar mechanism to N-terminal hydrolases
• Initial attack of amide by adjacent threonine is slow, rate limiting step
B-
BH+
H
H
O
O
O
CH3
H+
O
O
CH3
O
HN
N
H
R
CH3
R
R
O
H
NH2
H
O
H+
O
O
BH
O
OH
R
O
CH3
HO
O
NH2
O
R
CH3
O
H
O
H
NH2