Serine Proteases Components of the enzymatic pocket: Oxyanion hole Components of the enzymatic pocket: 1. Catalytic triad (asp-his-ser) 2. Oxyanion hole 3. Specificity pocket Specificity pocket Catalytic triad

Basic mechanism

Aspartyl Proteases Components of the enzymatic pocket: 1. Two aspartate residues 2. Enzyme bound water 3. Specificity for hydrophobic amino acids Charge repulsion of the two aspartate residues decreases side chain acidity

Basic mechanism -There is no enzyme-substrate intermediate!

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)

N-terminal hydrolases Examples include the proteasome, penicillin acylase, glutamine PPP amidohydrolase Catalytic domain consists of a bound water molecule and an N-terminal serine, threonine, or cysteine

Autoproteolysis Similar mechanism to N-terminal hydrolases Initial attack of amide by adjacent threonine is slow, rate limiting step