1.14.15.8 synthesis a steroid 15beta-hydroxylating whole-cell solvent tolerant biocatalyst is constructed by expressing the Bacillus megaterium steroid hydroxylase CYP106A2 in the solvent tolerant Pseudomonas putida S12. Testosterone hydroxylation is improved by a factor 16 by co-expressing Fer, a putative Fe-S protein from Bacillus subtilis. The specificity for 15beta-hydroxylation is improved by mutating threonine residue 248 of CYP106A2 into valine. These insights provide the basis for an optimized whole-cell steroid-hydroxylating biocatalyst that can be applied with an organic solvent phase 699068 1.14.15.8 synthesis an overexpressing Bacillus megaterium strain produces up to 115 mg/l/h 7beta-hydroxydehydroepiandrosterone in whole-cell conversions 745699 1.14.15.8 synthesis efficient approach towards the preparative scale synthesis of hydroxylated steroid derivatives. Improve CYP106A2-catalyzed steroid hydroxylation towards higher productivity and quantitative product formation. Because substrate transport into the cell limits the whole-cell biotransformation, activity can be increased sixfold by using membrane-free crude cell as biocatalyst 697322