SUBUNIT INTERACTION IN TRYPTOPHAN SYNTHASE OF ESCHERICHIA-COLI - CALORIMETRIC STUDIES ON ASSOCIATION OF ALPHA AND BETA-2 SUBUNITS

Association of the apo-β2 and the holo-(β-PLP)2 subunits of tryptophan synthase from Escherichia coli (l-seririe hydro-lyase (adding indole) (EC 4.2.1.20)) with a subunits of the same enzyme has been studied by microcalorirtletry. The results obtained from thermometric titrations clearly demonstrate that only the native complex α2β2 is formed, independent of an excess of α protein. The reaction of the holo-(β-PLP)2 withα subunits at 25°C is accompanied by a negative enthalpy change, which is almost twice as large as that for complex formation with the apo-β2 protein, thus indicating that the interaction enthalpy becomes more favorable in the presence of the coenzyme pyridoxal 5'-phosphate (PLP). Both reaction enthalpies show very large negative temperature coefficients, -3600± 100 cal K-1 (mol of β2)-1 being the value for the formation of the apoenzyme and -2300± 100 cal K-1 (mol of β2)-1 pertaining to formation of the holoenzyme. The studies on the association of αand β2 subunits in the two buffers revealed that at 25°C approximately 0.75 proton are absorbed in the presence and absence of the coenzyme, whereas at 35°C one proton is taken up from the solution when PLP is present, but two if the apo-β2 complex reacts. These results are a clear indication of energetic linkage between intersubunit interaction, hydrogen ion equilibria, and the binding of the coenzyme. © 1979, American Chemical Society. All rights reserved.