Testosterone 15beta-hydroxylation by solvent tolerant Pseudomonas putida S12.
A steroid 15beta-hydroxylating whole-cell solvent tolerant biocatalyst was constructed by expressing the Bacillus megaterium steroid hydroxylase CYP106A2 in the solvent tolerant Pseudomonas putida S12. Testosterone hydroxylation was improved by a factor 16 by co-expressing Fer, a putative Fe-S protein from Bacillus subtilis. In addition, the specificity for 15beta-hydroxylation was improved by mutating threonine residue 248 of CYP106A2 into valine. These new insights provide the basis for an optimized whole-cell steroid-hydroxylating biocatalyst that can be applied with an organic solvent phase.
Ruijssenaars
Harald J
HJ
TNO Quality of Life, Julianalaan 67, 2628BC Delft, The Netherlands. harald.ruijssenaars@tno.nl
Sperling
Eric M G M
EM
Wiegerinck
Peter H G
PH
Brands
Ferry T L
FT
Wery
Jan
J
de Bont
Jan A M
JA
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006900:Hydroxylation; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D030781:Organisms, Genetically Modified; D016958:Pseudomonas putida / Q000235:genetics*; D012997:Solvents / Q000494:pharmacology*; D013379:Substrate Specificity; D013739:Testosterone / Q000378:metabolism*
D001426:Bacterial Proteins; D012997:Solvents; D013739:Testosterone; D003577:Cytochrome P-450 Enzyme System; C517094:15beta-hydroxylase CYP106A2, Bacillus megaterium