Construction and in vivo assembly of a catalytically proficient and hyperthermostable de novo enzyme.

Autor:	Watkins DW; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Jenkins JMX; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Grayson KJ; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Wood N; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Steventon JW; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Le Vay KK; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Goodwin MI; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK.; School of Chemistry, University of Bristol, Bristol, BS8 1TS, UK., Mullen AS; Henry Wellcome Unit of Biological EPR, School of Chemistry, University of East Anglia, Norwich, NR4 7TJ, UK., Bailey HJ; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Crump MP; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK.; School of Chemistry, University of Bristol, Bristol, BS8 1TS, UK., MacMillan F; Henry Wellcome Unit of Biological EPR, School of Chemistry, University of East Anglia, Norwich, NR4 7TJ, UK., Mulholland AJ; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK.; School of Chemistry, University of Bristol, Bristol, BS8 1TS, UK., Cameron G; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK., Sessions RB; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK., Mann S; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK.; School of Chemistry, University of Bristol, Bristol, BS8 1TS, UK., Anderson JLR; School of Biochemistry, University of Bristol, University Walk, Bristol, BS8 1TD, UK. ross.anderson@bristol.ac.uk.; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Tyndall Avenue, Bristol, BS8 1TQ, UK. ross.anderson@bristol.ac.uk.
Jazyk:	angličtina
Zdroj:	Nature communications [Nat Commun] 2017 Aug 25; Vol. 8 (1), pp. 358. Date of Electronic Publication: 2017 Aug 25.
DOI:	10.1038/s41467-017-00541-4
Abstrakt:	Although catalytic mechanisms in natural enzymes are well understood, achieving the diverse palette of reaction chemistries in re-engineered native proteins has proved challenging. Wholesale modification of natural enzymes is potentially compromised by their intrinsic complexity, which often obscures the underlying principles governing biocatalytic efficiency. The maquette approach can circumvent this complexity by combining a robust de novo designed chassis with a design process that avoids atomistic mimicry of natural proteins. Here, we apply this method to the construction of a highly efficient, promiscuous, and thermostable artificial enzyme that catalyzes a diverse array of substrate oxidations coupled to the reduction of H 2 O 2 . The maquette exhibits kinetics that match and even surpass those of certain natural peroxidases, retains its activity at elevated temperature and in the presence of organic solvents, and provides a simple platform for interrogating catalytic intermediates common to natural heme-containing enzymes.Catalytic mechanisms of enzymes are well understood, but achieving diverse reaction chemistries in re-engineered proteins can be difficult. Here the authors show a highly efficient and thermostable artificial enzyme that catalyzes a diverse array of substrate oxidations coupled to the reduction of H 2 O 2 .
Databáze:	MEDLINE
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