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Free keywords:
tryptophan synthase; loop closure; pyroxidal phosphate; crystal structure; allosteric communication
Abstract:
The catalytic activity and substrate channeling of the pyridoxal 5′-phosphate-dependent tryptophan synthase α2β2 complex is regulated by allosteric interactions that modulate the switching of the enzyme between open, low activity and closed, high activity states during the catalytic cycle. The highly conserved αThr183 residue is part of loop αL6 and is located next to the α-active site and forms part of the α–β subunit interface. The role of the interactions of αThr183 in α-site catalysis and allosteric regulation was investigated by analyzing the kinetics and crystal structures of the isosteric mutant αThr183Val. The mutant displays strongly impaired allosteric α–β communication, and the catalytic activity of the α-reaction is reduced one hundred fold, whereas the β-activity is not affected. The structural work establishes that the basis for the missing inter-subunit signaling is the lack of loop αL6 closure even in the presence of the α-subunit ligands, 3-indolyl-d-glycerol 3′-phosphate, or 3-indolylpropanol 3′-phosphate. The structural basis for the reduced α-activity has its origins in the missing hydrogen bond between αThr183 and the catalytic residue, αAsp60.
