title: A Panel of TrpB Biocatalysts Derived from Tryptophan Synthase through the Transfer of Mutations that Mimic Allosteric Activation.
abstract: Naturally occurring enzyme homologues often display highly divergent activity with non-natural substrates. Exploiting this diversity with enzymes engineered for new or altered function, however, is laborious because the engineering must be replicated for each homologue. A small set of mutations of the tryptophan synthase β-subunit (TrpB) from Pyrococcus furiosus, which mimics the activation afforded by binding of the α-subunit, was demonstrated to have a similar activating effect in different TrpB homologues with as little as 57 % sequence identity. Kinetic and spectroscopic analyses indicate that the mutations function through the same mechanism: mimicry of α-subunit binding. From these enzymes, we identified a new TrpB catalyst that displays a remarkably broad activity profile in the synthesis of 5-substituted tryptophans. This demonstrates that allosteric activation can be recapitulated throughout a protein family to explore natural sequence diversity for desirable biocatalytic transformations.
authors: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, 91125, USA.
authors: Javier
authors: J
authors: Murciano-Calles
authors: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, 91125, USA.
authors: David K
authors: DK
authors: Romney
authors: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, 91125, USA.
authors: Sabine
authors: S
authors: Brinkmann-Chen
authors: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, 91125, USA.
authors: Andrew R
authors: AR
authors: Buller
authors: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, 91125, USA. frances@cheme.caltech.edu.
authors: Frances H
authors: FH
authors: Arnold
mesh_terms: D000494:Allosteric Regulation; D055162:Biocatalysis; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D015202:Protein Engineering; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020027:Pyrococcus furiosus / Q000201:enzymology; D014367:Tryptophan Synthase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D021122:Protein Subunits; D014367:Tryptophan Synthase
keywords: allostery; enzymes; protein engineering; tryptophan synthase