P450s for Pharmaceutical applications - a new whole cell biocatalyst by surface display of CYP102A1 via autodisplay

Schüürmann J, Lindhorst F, Jose J

Abstract

Whole cell biocatalysis offers significant advantages for industrial applications compared to purified enzymes, in particular, obsolete protein purification, enhanced enzyme stability andreusability. However, restriction to cell permeable substrates and products as well as cross reactions with intracellular enzymes could pose a severe challenge to specific applications. Onesolution is the presentation of enzymes on the cell surface to circumvent mass transfer problems and possible side reactions. Recently, our group demonstrated the surface display of activehuman CYP 3A4 using the Autodisplay technology5. The technique replaces the passenger domain of a native autotransporter by a peptide or protein of choice, which is then transported to thecell surface2. The current project focusses on the display of CYP102A1* from Bacillus megaterium for pharmaceutical applications. Surface exposure of the enzyme was confirmed by proteaseaccessibility tests, flow-cytometry analysis and activity measurements. CYP102A1, also known as BM3 is one of the best studied Cytochrome P450s to date and can be engineered to accept manypharmaceutical relevant substances. Thus, our whole cell biocatalyst can be used in the production of high value compounds, especially when mass transfer problems across the bacterialmembrane become a major issue.