Work

The CYP152-family P450 enzyme CypC of Bacillus subtilis converts non-natural substrates in plasma-driven biocatalysis

Year: 2025

Type: article

Abstract: Plasma-driven biocatalysis utilizes in situ H 2 O 2 production by atmospheric pressure plasmas to drive H 2 O 2 -dependent enzymatic reactions. Having previously established plasma-driven biocatalysis using recombinant unspecific peroxygenase from Agrocybe aegerita (r Aae UPO) to produce ( R )-1-phenylethanol from ethylbenzene, we here employed CypC from Bacillus subtilis 168 (synonyms: YbdT, P450BSβ), an integral enzyme of surfactin and fengycin biosynthesis. CypC naturally hydroxylates medium and long-chain carboxylic acids. With short-chain carboxylic acids as decoy molecules, it also converts non-natural substrates such as ethylbenzene. We optimized production and heme loading of CypC and established guaiacol and ABTS-based reactions to assess compatibility of CypC with plasma-driven biocatalysis regarding temperature and H 2 O 2 operating windows. With heptanoic acid as the decoy molecule and H 2 O 2 from stock solution, guaiacol and ABTS conversion yielded 18.28 and 21.13 nmol product min −1 nmol −1 CypC , respectively. We then supplied H 2 O 2 using a capillary plasma jet operated with 1280 ppm H 2 O in helium to convert ethylbenzene with immobilized CypC in a rotating bed reactor (5 ml reaction volume). After 120 min run time, a turnover number (TON) of 18.82 mol ( R )-1-PhOl mol −1 CypC was reached, demonstrating that plasma-driven biocatalysis can be extended to H 2 O 2 -dependent enzymes beyond r Aae UPO to expand the product range.

Source: Applied Microbiology and Biotechnology

DOI: 10.1007/s00253-025-13568-1