A Structural View of the Dissociation of Escherichia Coli Tryptophanase

Tryptophanase (Trpase) is a pyridoxal 5′-phosphate (PLP)-dependent homotetrameric enzyme which catalyzes the degradation of l-tryptophan. Trpase is known for its cold lability, i.e., a reversible loss of activity at low temperature (2°C), associated with tetramer dissociation. Escherichia coli Trpase dissociates into dimers, while Proteus vulgaris Trpase dissociates into monomers. Accordingly, Trpase is an appropriate model to study the quaternary structure of proteins. We aimed at understanding the differences in the mode of dissociation between the E. coli and P. vulgaris Trpases. In particular, the effect of mutations along the molecular axes of homotetrameric Trpase on its dissociation was studied. Two groups of mutants of the E. coli enzyme were created to resemble the amino-acid sequence of P. vulgaris Trpase. In one, residues 15 and 59 that are located along the molecular axis R (termed the noncatalytic axis) were mutated. The second group included a mutation at position 298, located along the molecular axis Q (termed the catalytic axis). Replacing amino-acid residues along the R axis resulted in dissociation of the tetramers into monomers, similar to the P. vulgaris Trpase, while replacing amino-acid residues along the Q axis resulted in dissociation into dimers only. The crystal structure of the V59M mutant of E. coli Trpase was also determined in its apo form and was found to be similar to that of the wild type. This study suggests that in E. coli Trpase hydrophobic interactions along the R axis hold the two monomers together more strongly, preventing the dissociation of the dimers into monomers. Mutation of position 298 along the Q axis to a charged residue resulted in tetramers that are less susceptible to dissociation. The results indicate that dissociation of E. coli Trpase into dimers occurs along the molecular Q axis.