title: Tryptophan Synthase Uses an Atypical Mechanism To Achieve Substrate Specificity.
abstract: Tryptophan synthase (TrpS) catalyzes the final steps in the biosynthesis of l-tryptophan from l-serine (Ser) and indole-3-glycerol phosphate (IGP). We report that native TrpS can also catalyze a productive reaction with l-threonine (Thr), leading to (2S,3S)-β-methyltryptophan. Surprisingly, β-substitution occurs in vitro with a 3.4-fold higher catalytic efficiency for Ser over Thr using saturating indole, despite a >82000-fold preference for Ser in direct competition using IGP. Structural data identify a novel product binding site, and kinetic experiments clarify the atypical mechanism of specificity: Thr binds efficiently but decreases the affinity for indole and disrupts the allosteric signaling that regulates the catalytic cycle.
authors.affiliation: Division of Chemistry and Chemical Engineering 210-41, California Institute of Technology , 1200 East California Boulevard, Pasadena, California 91125, United States.
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Buller
authors.affiliation: Division of Chemistry and Chemical Engineering 210-41, California Institute of Technology , 1200 East California Boulevard, Pasadena, California 91125, United States.
authors.forename: Paul
authors.initials: P
authors.lastname: van Roye
authors.affiliation: Division of Chemistry and Chemical Engineering 210-41, California Institute of Technology , 1200 East California Boulevard, Pasadena, California 91125, United States.
authors.forename: Javier
authors.initials: J
authors.lastname: Murciano-Calles
authors.affiliation: Division of Chemistry and Chemical Engineering 210-41, California Institute of Technology , 1200 East California Boulevard, Pasadena, California 91125, United States.
authors.forename: Frances H
authors.identifier: http://orcid.org/0000-0002-4027-364X
authors.initials: FH
authors.lastname: Arnold
mesh_terms: D000494:Allosteric Regulation; D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D055162:Biocatalysis; D053898:Biosynthetic Pathways; D018360:Crystallography, X-Ray; D005994:Glycerophosphates / Q000737:chemistry / Q000378:metabolism*; D007211:Indoles / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D011485:Protein Binding; D000072417:Protein Domains; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D020027:Pyrococcus furiosus / Q000201:enzymology; D012694:Serine / Q000737:chemistry / Q000378:metabolism*; D013053:Spectrophotometry; D013379:Substrate Specificity; D013912:Threonine / Q000737:chemistry / Q000378:metabolism; D014364:Tryptophan / Q000031:analogs & derivatives / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014367:Tryptophan Synthase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; D005994:Glycerophosphates; D007211:Indoles; D021122:Protein Subunits; C538978:beta-methyltryptophan; D013912:Threonine; C008117:indoleglycerol phosphate; D012694:Serine; C030374:indole; D014364:Tryptophan; D014367:Tryptophan Synthase