Pressure effects on the association of the and 2 subunits of tryptophan synthetase from Escherichia coli and Salmonella typhimurium

Sucrose density gradient centrifugation was employed to study the association of the α and β₂ subunits of the enzyme tryptophan synthetase from Escherichia coli and Salmonella typhimurium. In both cases, the fully associated enzyme (α₂β₂) showed a sedimentation coefficient of 6.4 S, in agreement with the values reported by other workers for the E. coli enzyme. The substrate, l-serine, and the cofactor, pyridoxal phosphate, were required for complex formation in both cases. Generation of moderately high pressures by increasing the centrifuge speed from 39,000 rpm to 50,000 rpm was found to interfere with complex formation in both species at 5 °C. This effect was reversed by a temperature increase from 5 °C to 20 °C or by low concentrations of a nonpolar solvent, ethanol, at 5 °C. These results indicate that hydrophobic bonding plays an important role in the formation of the active tryptophan synthetase α₂β₂ complex. Monovalent and divalent cations also interfered with the formation of the α₂β₂ complex, indicating the possibility that ionic bonds are also involved.