2j9z

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spinqualitylabelsall models 2j9z, resolution 1.80Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

TRYPTOPHAN SYNTHASE T110 MUTANT COMPLEX

Overview

In the PLP-requiring alpha2beta2 tryptophan synthase complex, recognition, of the substrate l-Ser at the beta-site includes a loop structure, (residues beta110-115) extensively H-bonded to the substrate, alpha-carboxylate. To investigate the relationship of this subsite to, catalytic function and to the regulation of substrate channeling, two loop, mutants were constructed: betaThr110 --> Val, and betaGln114 --> Asn. The, betaT110V mutation greatly impairs both catalytic activity in the, beta-reaction, and allosteric communication between the alpha- and, beta-sites. The crystal structure of the betaT110V mutant shows that the, modified l-Ser carboxylate subsite has altered protein interactions that, impair beta-site catalysis and the communication of allosteric signals, between the alpha- and beta-sites. Purified betaQ114N consists of two, species of mutant protein, one with a reddish color (lambdamax = 506 nm)., The reddish species is unable to react with l-Ser. The second betaQ114N, species displays significant catalytic activities; however, intermediates, obtained on reaction with substrate l-Ser and substrate analogues exhibit, perturbed UV/vis absorption spectra. Incubation with l-Ser results in the, formation of an inactive species during the first 15 min with lambdamax, approximately 320 nm, followed by a slower conversion over 24 h to the, species with lambdamax = 506 nm. The 320 and 506 nm species originate from, conversion of the alpha-aminoacrylate external aldimine to the internal, aldimine and alpha-aminoacrylate, followed by the nucleophilic attack of, alpha-aminoacrylate on C-4' of the internal aldimine to give a covalent, adduct with PLP. Subsequent treatment with sodium hydroxide releases a, modified coenzyme consisting of a vinylglyoxylic acid moiety linked, through C-4' to the 4-position of the pyridine ring. We conclude that the, shortening of the side chain accompanying the replacement of beta114-Gln, by Asn relaxes the steric constraints that prevent this reaction in the, wild-type enzyme. This study reveals a new layer of structure-function, interactions essential for reaction specificity in tryptophan synthase.

About this Structure

2J9Z is a Protein complex structure of sequences from Salmonella typhimurium with and as ligands. Active as Tryptophan synthase, with EC number 4.2.1.20 Known structural/functional Sites: and . Full crystallographic information is available from OCA.

Reference

betaQ114N and betaT110V Mutations Reveal a Critically Important Role of the Substrate alpha-Carboxylate Site in the Reaction Specificity of Tryptophan Synthase., Blumenstein L, Domratcheva T, Niks D, Ngo H, Seidel R, Dunn MF, Schlichting I, Biochemistry. 2007 Nov 16;. PMID:18004874

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