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.
MeSH terms
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Allosteric Regulation
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Archaeal Proteins / chemistry
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Archaeal Proteins / metabolism*
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Binding Sites
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Biocatalysis
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Biosynthetic Pathways*
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Crystallography, X-Ray
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Glycerophosphates / chemistry
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Glycerophosphates / metabolism*
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Indoles / chemistry
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Indoles / metabolism
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Kinetics
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Models, Molecular
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Molecular Structure
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Protein Binding
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Protein Domains
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Protein Subunits / chemistry
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Protein Subunits / metabolism
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Pyrococcus furiosus / enzymology
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Serine / chemistry
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Serine / metabolism*
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Spectrophotometry
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Substrate Specificity
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Threonine / chemistry
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Threonine / metabolism
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Tryptophan / analogs & derivatives
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Tryptophan / biosynthesis*
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Tryptophan / chemistry
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Tryptophan / genetics
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Tryptophan / metabolism
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Tryptophan Synthase / chemistry
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Tryptophan Synthase / metabolism*
Substances
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Archaeal Proteins
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Glycerophosphates
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Indoles
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Protein Subunits
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beta-methyltryptophan
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Threonine
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indoleglycerol phosphate
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Serine
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indole
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Tryptophan
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Tryptophan Synthase