Abstract
The catalytic activity of the pyridoxal 5'-phosphate-dependent tryptophan synthase alpha(2)beta(2) complex is allosterically regulated. The hydrogen bond between the helix betaH6 residue betaSer(178) and the loop alphaL6 residue Gly(181) was shown to be critical in ligand-induced intersubunit signaling, with the alpha-beta communication being completely lost in the mutant betaSer(178) --> Pro (Marabotti, A., De Biase, D., Tramonti, A., Bettati, S., and Mozzarelli, A. (2001) J. Biol. Chem. 276, 17747-17753). The structural basis of the impaired allosteric regulation was investigated by determining the crystal structures of the mutant betaSer(178) --> Pro in the absence and presence of the alpha-subunit ligands indole-3-acetylglycine and glycerol 3-phosphate. The mutation causes local and distant conformational changes especially in the beta-subunit. The ligand-free structure exhibits larger differences at the N-terminal part of helix betaH6, whereas the enzyme ligand complexes show differences at the C-terminal side. In contrast to the wild-type enzyme loop alphaL6 remains in an open conformation even in the presence of alpha-ligands. This effects the equilibrium between active and inactive conformations of the alpha-active site, altering k(cat) and K(m), and forms the structural basis for the missing allosteric communication between the alpha- and beta-subunits.
Highlights
The catalytic activity of the pyridoxal 5-phosphatedependent tryptophan synthase ␣22 complex is allosterically regulated
The hydrogen bond between the helix H6 residue Ser178 and the loop ␣L6 residue Gly181 was shown to be critical in ligand-induced intersubunit signaling, with the ␣- communication being completely lost in the mutant Ser178 3 Pro (Marabotti, A., De Biase, D., Tramonti, A., Bettati, S., and Mozzarelli, A. (2001) J
The hydrogen bond between the carboxylate of the active conformation of ␣Glu49 and of IAG has been observed in the TRPSIAG complex [24]
Summary
The catalytic activity of the pyridoxal 5-phosphatedependent tryptophan synthase ␣22 complex is allosterically regulated. The structural basis of the impaired allosteric regulation was investigated by determining the crystal structures of the mutant Ser178 Pro in the absence and presence of the ␣-subunit ligands indole-3-acetylglycine and glycerol 3-phosphate. In contrast to the wild-type enzyme loop ␣L6 remains in an open conformation even in the presence of ␣-ligands. This effects the equilibrium between active and inactive conformations of the ␣-active site, altering kcat and Km, and forms the structural basis for the missing allosteric communication between the ␣- and -subunits. The ␣22 complex of tryptophan synthase (TRPS) (EC 4.2.1.20) is a bifunctional enzyme that is considered a para-
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