THERMALLY-INDUCED UNFOLDING OF THE TRYPTOPHAN SYNTHASE ALPHA(2)BETA-2 MULTIENZYME COMPLEX FROM SALMONELLA-TYPHIMURIUM

Tryptophan synthase (TS) from Salmonella typhimurium (144,000 M(r)) is a bifunctional enzyme composed of two heterodimers arranged in an extended alpha beta beta alpha geometry [Hyde et al., J. Biol. Chem. 263, 17857, 1988]. The thermal unfolding of TS alpha(2) beta(2) and isolated alpha and beta subunits has been studied by differential scanning calorimetry (DSC), circular dichroism, and UV spectroscopy. DSC profiles of the holo-alpha(2) beta(2) complex containing pyridoxal phosphate (PLP) bound to each beta chain are characterized by two well-resolved endotherms centered at approximate to 52 and approximate to 81 degrees C. Conformational transitions in beta chains (T-m approximate to 46 degrees C) as well as sequential unfolding reactions in a chains contribute to the low-temperature endotherm whereas major unfolding of the beta dimer occurs between 74 and 82 degrees C. The thermally induced unfolding disrupts approximate to 70% of the secondary structure with a total enthalpy change of 3600 +/- 100 kJ mol(-1) for holo-alpha(2) beta(2). Stabilizing interactions between alpha and beta chains are apparent. The cofactor PLP not only markedly stabilizes beta chains in beta(2) but also increases the cooperativity of unfolding beta chains. Calorimetric and spectral measurements suggest a sequential unfolding pathway for the tryptophan synthase alpha(2) beta(2) complex. Thermodynamic results are correlated with available structural information.