Back to Search
Start Over
Production of 14α-Hydroxy Progesterone Using a Steroidal Hydroxylase from Cochliobolus lunatus Expressed in Escherichia coli.
- Source :
-
Catalysts (2073-4344) . Apr2024, Vol. 14 Issue 4, p247. 14p. - Publication Year :
- 2024
-
Abstract
- Steroids with hydroxylation at C14 are drawing increased attention because of their diverse biological activities and applications. P-450lun from Cochliobolus lunatus is the first fungal cytochrome P450 reported to have 14α-hydroxylase activity. Studies have shown that P-450lun catalyzes the hydroxylation of progesterone (PROG) at C14α with low regiospecificity and activity. To improve its regiospecificity and activity for PROG, truncated forms of P-450lun and its cognate redox partner CPRlun were functionally co-expressed in Escherichia coli. Then, a semi-rational protein engineering approach was applied to P-450lun, resulting in a double-site mutant E109A/F297W with enhanced 14α-position selectivity for PROG compared with the wild-type P-450lun (97% vs. 28%). Protein structure analysis revealed that the F297W substitution can hinder the binding pose for 11β-hydroxylation product formation. Finally, whole-cell catalysis was optimized, and the final titer of 14α-OH-PROG reached 16.0 mg/L. This is the first report where a fungal 14α-hydroxylase was functionally expressed in Escherichia coli. The steroid hydroxylation system obtained in this study can serve as a basis for the synthesis of 14α-hydroxylated PROG and the rapid evolution of eukaryotic cytochrome P-450lun. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 20734344
- Volume :
- 14
- Issue :
- 4
- Database :
- Academic Search Index
- Journal :
- Catalysts (2073-4344)
- Publication Type :
- Academic Journal
- Accession number :
- 176874590
- Full Text :
- https://doi.org/10.3390/catal14040247