Expanding the Substrate Scope of Nitrating Cytochrome P450 TxtE by Active Site Engineering of a Reductase Fusion

Rakesh Saroay, Gheorghe Doru Roiban, Lona M. Alkhalaf, Gregory L. Challis

Research output: Contribution to journalArticleResearchpeer-review

12 Citations (Scopus)

Abstract

Aromatic nitration reactions are a cornerstone of organic chemistry, but are challenging to scale due to corrosive reagents and elevated temperatures. The cytochrome P450 TxtE nitrates the indole 4-position of l-tryptophan at room temperature using NO, O2 and NADPH, and has potential to be developed into a useful aromatic nitration biocatalyst. However, its narrow substrate scope (requiring both the α-amino acid and indole functionalities) have hindered this. Screening of an R59 mutant library of a TxtE-reductase fusion protein identified a variant (R59C) that nitrates tryptamine, which is not accepted by native TxtE. This variant exhibits a broader substrate scope than the wild type enzyme and is able to nitrate a range of tryptamine analogues, with significant alterations to the aromatic and aminoethyl moieties.

Original languageEnglish
Pages (from-to)2262-2265
Number of pages4
JournalChemBioChem
Volume22
Issue number13
DOIs
Publication statusPublished - 1 Jul 2021

Keywords

  • Biocatalysis
  • Indole
  • Nitration
  • Protein Engineering
  • ARC Centre of Excellence for Innovations in Peptide and Protein Science

    Craik, D. J. (Primary Chief Investigator (PCI)), Payne, R. J. (Chief Investigator (CI)), Belov, K. (Chief Investigator (CI)), Jolliffe, K. A. (Chief Investigator (CI)), New, E. (Chief Investigator (CI)), Fairlie, D. P. (Chief Investigator (CI)), King, G. F. (Chief Investigator (CI)), Henriques, S. T. (Chief Investigator (CI)), Otting, G. (Chief Investigator (CI)), Malins, L. (Chief Investigator (CI)), Jackson, C. (Chief Investigator (CI)), Cryle, M. (Chief Investigator (CI)), Challis, G. (Chief Investigator (CI)), Colgrave, M. (Chief Investigator (CI)), Ploegh, H. (Partner Investigator (PI)), Baker, D. A. (Partner Investigator (PI)), Suga, H. (Partner Investigator (PI)), Davis, B. G. (Partner Investigator (PI)), Pentelute, B. (Partner Investigator (PI)), van der Donk, W. (Partner Investigator (PI)), Tavassoli, A. (Partner Investigator (PI)) & Savage, G. (Partner Investigator (PI))

    23/12/2021/01/28

    Project: Research

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