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B6db references: 26553994

type Journal Article
authors Buller AR, Brinkmann-Chen S, Romney DK, Herger M, Murciano-Calles J, Arnold FH
title Directed evolution of the tryptophan synthase β-subunit for stand-alone function recapitulates allosteric activation
journal Proc Natl Acad Sci U S A
sel selected
ui 26553994
year (2015)
volume 112
number 47
pages 14599-604
keywords PLP; allostery; noncanonical amino acid; protein engineering
abstract Enzymes in heteromeric, allosterically regulated complexes catalyze a rich array of chemical reactions. Separating the subunits of such complexes, however, often severely attenuates their catalytic activities, because they can no longer be activated by their protein partners. We used directed evolution to explore allosteric regulation as a source of latent catalytic potential using the β-subunit of tryptophan synthase from Pyrococcus furiosus (PfTrpB). As part of its native αββα complex, TrpB efficiently produces tryptophan and tryptophan analogs; activity drops considerably when it is used as a stand-alone catalyst without the α-subunit. Kinetic, spectroscopic, and X-ray crystallographic data show that this lost activity can be recovered by mutations that reproduce the effects of complexation with the α-subunit. The engineered PfTrpB is a powerful platform for production of Trp analogs and for further directed evolution to expand substrate and reaction scope.
last changed 2018/01/31 10:40

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