Optimisation and scale-up of a-glucuronidase production by recombinant Saccharomyces cerevisiae in aerobic fed-batch culture with constant growth rate

摘要

α-Glucuronidase (EC 3.2.1.139) of family GH 115 from Scheffersomyces stipitis is a valuable enzyme for the modification of water-soluble xylan into insoluble biopolymers, due to its unique ability to act on polymeric xylans. The influence of growth rate on the production of a-glucuronidase by recombinant Saccharomyces cerevisiae MH1000pbkl OD-glu in glucose-limited fed-batch culture was studied at 14 and 100L scale. At and below the critical specific growth rate (μ_(crit)) of 0.12h~(-1)H at 14L scale, the biomass yield coefficient (Y_(x/s)) remained constant at 0.4 gg~(-1) with no ethanol production, whereas ethanol yields relative to biomass (k_(eth/x)) of up to 0.54 gg~(-1) and a steady decrease in Y_(x/s) were observed at μ>0.12h~(-1). Production of a-glucuronidase was growth associated at a product yield (k(α-glu/x)) of 0.45 mgg~(-1), with the highest biomass (37.35 g/L) and a-glucuronidase (14.03 mg/L) concentrations, were recorded during fed-batch culture at or near to μ_(crit). Scale-up with constant k_la from 14 to 100 L resulted in ethanol concentrations of up to 2.5 g/L at μ= 0.12 h~(-1). At this scale, a-glucuronidase yield could be maximised at growth rates below μ_(crit), to prevent localised high glucose concentration pockets at the feed entry zone that would induce oxido-reductive metabolism. This is the first report where recombinant production of a-glucuronidase (EC 3.2.1.139) by S. cerevisiae was optimised for application at pilot scale.