The α2β2 tryptophan synthase complex is a model enzyme for understanding allosteric regulation. We report the functional and regulatory properties of the βS178P mutant. Ser-178 is located at the end of helix 6 of the β subunit, belonging to the domain involved in intersubunit signaling. The carbonyl group of βSer-178 is hydrogen bonded to Gly-181 of loop 6 of the α subunit only when α subunit ligands are bound. An analysis by molecular modeling of the structural effects caused by the βS178P mutation suggests that the hydrogen bond involving αGly-181 is disrupted as a result of localized structural perturbations. The ratio of α to β subunit concentrations was calculated to be 0.7, as for the wild type, indicating the maintenance of a tight α-β complex. Both the activity of the α subunit and the inhibitory effect of the α subunit ligands indole-3-acetylglycine and D,L-α -glycerol-3-phosphate were found to be the same for the mutant and wild type enzyme, whereas the β subunit activity of the mutant exhibited a 2-fold decrease. In striking contrast to that observed for the wild type, the allosteric effectors indole-3-acetylglycine and D,L-α -glycerol-3-phosphate do not affect the β activity. Accordingly, the distribution of L-serine intermediates at the β-site, dominated by the α-aminoacrylate, is only slightly influenced by α subunit ligands. Binding of sodium ions is weaker in the mutant than in the wild type and leads to a limited increase of the amount of the external aldimine intermediate, even at high pH, whereas binding of cesium ions exhibits the same affinity and effects as in the wild type, leading to an increase of the α -aminoacrylate tautomer absorbing at 450 nm. Crystals of the βS178P mutant were grown, and their functional and regulatory properties were investigated by polarized absorption microspectrophotometry. These findings indicate that (i) the reciprocal activation of the α and β activity in the α2β2 complex with respect to the isolated subunits results from interactions that involve residues different from βSer-178 and (ii) βSer-178 is a critical residue in ligand-triggered signals between α and β active sites.
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