Plasma-driven biocatalysis using the cytochrome P450 enzyme CYP152 _BSβ

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 (ETBE), we here employed CYP152 from Bacillus subtilis (CYP152 _BSβ ). CYP152 _BSβ naturally hydroxylates medium and long-chain carboxylic acids, and, with short-chain carboxylic acids as decoy molecules, also converts non-natural substrates such as ETBE. To produce active CYP152 _BSβ overexpression and heme loading were optimized. The conversion of the non-natural substrates guaiacol and ABTS with heptanoic acid as decoy molecule and H ₂ O ₂ from stock solution yielded 18.28 and 21.13 nmol product min ^-1 , respectively. These reactions also served to assess compatibility of CYP152 _BSβ with plasma-driven biocatalysis regarding temperature and H ₂ O ₂ operating windows. To establish CYP152 _BSβ -based plasma-driven biocatalysis, immobilized enzyme in a rotating bed reactor (5 ml reaction volume) was then supplied with H ₂ O ₂ from a capillary plasma jet operated with 1280 ppm H ₂ O in helium. After a 120 min run time a turnover number (TON) of 18.82 mol _{( R )-1-PhOl} was reached. We conclude that plasma-driven biocatalysis can be extended to other H ₂ O ₂ -dependent enzymes. Future efforts will be directed at increasing the TON and product range.