lytic performance of recombinant P450-containing resting cells, (ii) lyophilized recombinant E. coli cells is often applied for P450-mediated biocatalysis, when (iii) metabolism-independent regeneration of NAD(P)H is ensured. The use of these procedures illustrates interesting perspectives for practical applications of cytochrome P450s for singleor multi-step reactions.Abbreviations CYP: Cytochrome P450, Pdr:putidaredoxin reductase from Pseudomonas putida.; Pdx: Putidaredoxin from Pseudomonas putida.; Re-ADH: Alcohol dehydrogenase from Rhodococcus erythropolis.; NADH: Nicotinamide adenine dinucleotide..Supplementary InformationThe online version consists of supplementary material offered at doi. org/10.1186/s13568-021-01319-0. Extra file 1: Table S1: Synthetic oligonucleotides for cloning. Table S2: Plasmids utilised in this study. Table S3: Analysis of testosterone 1 and metabolites 2-10 that have been formed for the duration of CYP105D-mediated oxidation. Figure S1: Impact of freezing and glycerol addition in the course of lyophilization on conversion catalyzed by E. coli C43 (DE3) pET22b-cyp105D + pCOLADuet- pdx-pdr-adh. E. coli cells have been after or twice frozen at – 80 and after that lyophilized for either 24 h (black) or 48 h (grey). Figure S2: Exemplary LC/MS-chromatogram displaying the oxidation of testosterone 1 to the goods 2-10 by the CYP105D-based E. coli whole-cell biocatalyst (pink) in comparison to a negative manage (black). Figure S3: SDS-PAGE analysis of E. coli C43 (DE3) strains for whole-cell biocatalysis. Figure S4: Impact of NADH addition on testosterone 1 conversion mediated by the lyophilized whole-cell catalyst with out ADH. NADH was added up to four occasions (number in brackets) every 2 h. Acknowledgements We thank Sebastian H zel for technical help. Authors’ contributions TH planned, designed and carried out most experiments, analyzed all of the data, and drafted the manuscript. AR performed and evaluated experiments with different preparations of P450 whole-cell catalysts. LMW contributed in initial experimental design and style with regard to building of expression vectors, gene expression and activity measurements. VBU gave advices inside the investigation work, helped in drafting the manuscript, and revised the manuscript. All authors study and authorized the final manuscript. Funding Open Access funding enabled and organized by Projekt DEAL. Economic support was kindly supplied by the Federal Ministry of Education and Analysis [Grant Quantity 031A223A] beneath the umbrella of your ERA-IB2 3rd get in touch with project `HyPerIn’ [Project Number EIB.12.026]. Availability of information and supplies All data generated or analyzed for the duration of this study are incorporated in this published report and its More files.DeclarationsEthics approval and consent to participate Not applicable. Consent for publication Not applicable. CCR8 Agonist Compound competing interests The authors declare no competing interests. Author details 1 Institute of Biochemistry, Heinrich-Heine University D CYP3 Inhibitor Biological Activity seldorf, Universit sstra 1, 40225 D seldorf, Germany. 2 Present Address: Division of Biotechnology, Delft University of Technologies, van der Maasweg 9, 2629HZ Delft, The Netherlands. three Present Address: School of Chemistry, University of Southampton, B30, University Road, SO17 1BJ Southampton, UK. Received: 12 September 2021 Accepted: 16 NovemberHilberath et al. AMB Express(2021) 11:Page 10 ofReferences Abokitse K, Hummel W (2003) Cloning, sequence analysis, and heterologous expression of your gene encoding a (S)-specific alcohol dehydrogen