Using Pseudomonas putida with surface displayed cellulases as whole cell biocatalyst for degradation of lignocellulosic biomass

Lenz F, Tozakidis IEP, Brossette T, Maas RM, Jose J

Abstract

Lignocellulose is the most abundant polymer on earth and often waste product of agricultural and paper industry. Its sustainable large scale degradation by cellulases to glucose is hindered for economic reasons, whereby the production of the enzymes causes the highest costs [1]. To simplify this process, we used the Gram-negative bacterium PseudomonasputidaKT2440 for surface presentation of endocellulase CelA, exocellulase CelK and β-glucosidase BglA from the thermophilic bacterium Ruminiclostridiumthermocellum as MaximizedAutoTransporter mediated Expression (MATE) fusion proteins. The costs for enzyme production could be reduced to the costs for cell cultivation and the immobilized enzymes could be regained by separation of the whole cells [2]. Cells displaying each one of the cellulases showed activity with their substrates carboxymethylcellulose (15,3 mU/mLOD1), 4-nitrophenyl-β-D-cellobioside (37,1 mU/mLOD1) and 4-nitrophenyl-β-D-glucopyranoside (0,5mU/mLOD1). After five repeated reaction cycles, the whole cell biocatalysts retained more than 75% of their initial activity. Also after 24 hours of preincubation at 55 oC, the cells still showed more than 75% activity. Finally, the combination of cells expressing MATE-CelA, cells expressing MATE-CelK and cells expressing MATE-BglA released 20 µg glucose per ml cell suspension in 24 hours [3].