Serine Protease Inhibitors - Conformational Change and Inhibitory Mechanism

Conformational Change and Inhibitory Mechanism

Early studies on serpins revealed that the mechanism by which these molecules inhibit target proteases appeared distinct from the lock-and-key-type mechanism utilised by small protease inhibitors such as the Kunitz-type inhibitors (e.g. basic pancreatic trypsin inhibitor). Indeed, serpins form covalent complexes with target proteases. Structural studies on serpins also revealed that inhibitory members of the family undergo an unusual conformational change, termed the Stressed to Relaxed (S to R) transition. During this structural transition the RCL inserts into β-sheet A (in red in figure 4 and 5) and forms an extra (fourth) β-strand. The serpin conformational change is key to the mechanism of inhibition of target proteases.

When attacking a substrate, serine proteases catalyze peptide bond cleavage in a two-step process. Initially, the catalytic serine performs a nucleophilic attack on the peptide bond of the substrate (Figure 5). This releases the new N-terminus and forms an ester-bond between the enzyme and the substrate. This covalent enzyme-substrate complex is called an acyl enzyme intermediate. Subsequent to this, this ester bond is hydrolysed and the new C-terminus is released. The RCL of a serpin acts as a substrate for its cognate protease. However, after the RCL is cleaved, but prior to hydrolysis of the acyl-enzyme intermediate, the serpin rapidly undergoes the S-to-R transition. Since the RCL is still covalently attached to the protease via the ester bond, the S-to-R transition causes the protease to be moved from the top to the bottom of the serpin. At the same time, the protease is distorted into a conformation, where the acyl enzyme intermediate is hydrolysed extremely slowly. The protease thus remains covalently attached to the target protease and is thereby inhibited. Further, since the serpin has to be cleaved to inhibit the target protases, inhibition consumes the serpin as well. Serpins are therefore irreversible enzyme inhibitors. The serpin mechanism of inhibition is illustrated in figures 5 and 6, and several movies illustrating the serpin mechanism can be viewed at this link.