Co-factor regeneration via Autodisplay – Towards industrial applications with surface displayed enzymes

Schüürmann J, Lang K, Jose J

Abstract

Costly purification steps and stability problems often renders the use of enzymes inefficient for industrial applications. Whole cell biocatalysts overcome these limitations and thus arepredestined for the application in biotechnological processes. However, restriction to cell permeable substrates and products as well as cross reactions with intracellular enzymes poses a severechallenge to specific applications. One solution is the presentation of enzymes on the cell surface to circumvent mass transfer problems and possible side reactions. The Autodisplay techniquecan be used to display enzymes on the cell surface of E. coli by replacing the passenger domain of a native autotransporter. Bringing an enzyme to the cell surface blocks the access tointracellular co-factors, which are too expensive to be added stoichiometrically. To keep the advantages of surface displayed catalysts we used the Autodisplay technology to present variousdehydrogenases on the cell surface of E. coli to regenerate NADPH. Surface display was confirmed by protease accessibility tests and FACS analysis and NADPH production was measuredphotometrically. We first concentrated on an formate dehydrogenase, but the efficiency of the system was limited by the number of enzymes on the bacterial surface and high amounts of wholecellcatalysts were needed for effective production of NADPH. Thus, we are currently exploring alternative enzymes with higher specific activities for cell associated co-factor regeneration.