Genetic Code Expansion in Shewanella oneidensis MR-1 Allows Site-Specific Incorporation of Bioorthogonal Functional Groups into a c -Type Cytochrome.

Autor:	Lockwood CWJ; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Nash BW; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Newton-Payne SE; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., van Wonderen JH; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Whiting KPS; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Connolly A; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Sutton-Cook AL; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Crook A; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Aithal AR; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Edwards MJ; School of Life Sciences, University of Essex, Colchester CO4 3SQ, U.K., Clarke TA; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Sachdeva A; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Butt JN; School of Chemistry and School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K.
Jazyk:	angličtina
Zdroj:	ACS synthetic biology [ACS Synth Biol] 2024 Sep 20; Vol. 13 (9), pp. 2833-2843. Date of Electronic Publication: 2024 Aug 19.
DOI:	10.1021/acssynbio.4c00248
Abstrakt:	Genetic code expansion has enabled cellular synthesis of proteins containing unique chemical functional groups to allow the understanding and modulation of biological systems and engineer new biotechnology. Here, we report the development of efficient methods for site-specific incorporation of structurally diverse noncanonical amino acids (ncAAs) into proteins expressed in the electroactive bacterium Shewanella oneidensis MR-1. We demonstrate that the biosynthetic machinery for ncAA incorporation is compatible and orthogonal to the endogenous pathways of S. oneidensis MR-1 for protein synthesis, maturation of c -type cytochromes, and protein secretion. This allowed the efficient synthesis of a c -type cytochrome, MtrC, containing site-specifically incorporated ncAA in S. oneidensis MR-1 cells. We demonstrate that site-specific replacement of surface residues in MtrC with ncAAs does not influence its three-dimensional structure and redox properties. We also demonstrate that site-specifically incorporated bioorthogonal functional groups could be used for efficient site-selective labeling of MtrC with fluorophores. These synthetic biology developments pave the way to expand the chemical repertoire of designer proteins expressed in S. oneidensis MR-1.
Databáze:	MEDLINE
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