Hydrostatic Pressure as a Structural and Mechanistic Probe of Tryptophan Synthase and Tryptophan Indole‐lyase

Abstract

The effects of hydrostatic pressure on the conformation and activity of tryptophan (Trp) synthase α2β2 complex from Salmonella typhimurium and tryptophan indole-lyase (Trpase) from Escherichia coli have been examined. The data demonstrate that the open conformation of Trp synthase is favored by higher pressure. The ΔVo and Ko for the equilibrium between the closed and open conformations of the Trp synthase-L-Ser complex are −126 mL/mol and 0.12 for the Na+form and −171 mL/mol and 2.3 x 10−4 for the NH4+ form. Pressure-jump experiments of the H463F mutant Trpase L-Trp and L-Met complexes show relaxations at 500 nm after rapid pressure changes of 100–400 bar for the complexes with both L-Trp and L-Met. However, the apparent rate constants for relaxation of L-Trp, but not L-Met, show a significant increase with pressure. These results suggest that there may be a contribution to the deprotonation reaction of Trpase either from quantum mechanical tunneling or from a mechanical coupling of protein motion and proton transfer for the reaction of L-Trp, but not with L-Met.