Proteases (also proteinases, peptidases) encompass a wide array of enzymes that catalyse the cleavage of peptide bond. There are four main classes of peptidases: cysteine proteases, aspartate proteases, metalloproteases and serine proteases. They are classified on the basis of the species involved in the catalysis. Serine proteases contain a highly reactive serine residue directly participating in catalysis. They comprise of nearly one third of all peptidases known and are implicated in many important processes such as digestion, blood clotting and immune response. Naturally occurring breakdown of the peptide bond is an extremely slow process. This is due to the high stability of the peptide bond (10-1000 years at neutral pH). Proteases adapt a common strategy to facilitate proteolysis. The mechanism requires three chemical features of the active site:
In serine proteases the nucleophile is an unusually reactive serine residue, serine 195. The unusual reactivity of serine 195 is due to the charge relay system (also known as the catalytic triad):
On binding of the substrate to chymotrypsin, His 57 attracts the proton from the OH group of Ser 195 which then launches a nucleophilic attack on the terminal carbon atom (step 2). A tetrahedral intermediate is formed and stabilised by hydrogen bonding of oxygen with the side chain of Gly 193 and the main chain NH group of Ser 195 . The amide accepts a proton from His 57 and dissociates (step 3). Following the breakdown of the peptide bond an acyl-enzyme intermediate is formed (step 4).
His 57 accepts a proton from water which displaces the amine. The resulting –OH ion attacks the carbon atom on the acyl group (Step 5). The second tetrahedral intermediate is created and stabilised by the same hydrogen bonds as previously (Gly 193 and Ser 195). The oxygen on the Ser 195 side chain accepts a proton from His 57 (step 6). The acyl group dissociates and the catalytic triad is restored (step 1).
The shape and properties of the enzyme’s binding pocket account for its specificity:
Interestingly the replacement of Asp 189 by serine partially converts trypsin to chymotrypsin.
Subtilisin is a bacterial serine protease secreted by Bacillus subtilis. The enzyme contains the same catalytic triad as chymotrypsin. However, the overall primary and secondary structures of both proteases are different due to their separate evolutionary origin. The relationship of subtilisin and the eukaryotic proteases is an example of convergent evolution.
Serine proteases encompass a variety of enzymes involved in:
Serine proteinases are inactivated by serpins (Serine Protease Inhibitors). Regulation of peptidases is often important to counterbalance their destructive activity. For instance α-1-antitrypsin serpin inhibits the neutrophil elastase. Serpins bind to the active site of the proteolytic enzyme forming a covalent serpin-proteinase complex. The conformational change leads to the partial unfolding of the substrate and halts its activity.
Mechanism of chymotrypsin animation:
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