Optimization of norovirus virus-like particle production inPichia pastorisusing a real-time near-infrared bioprocess monitor

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The production of norovirus virus-like particles (NoV VLPs) displaying NY-ESO-1 cancer testis antigen in Pichia pastoris BG11 Mut+ has been enhanced through feed-strategy optimization using a near-infrared bioprocess monitor (RTBio® Bioprocess Monitor, ASL Analytical, Inc.), capable of monitoring and controlling the concentrations of glycerol and methanol in real-time. The production of NoV VLPs displaying NY-ESO-1 in P. pastoris has potential as a novel cancer vaccine platform. Optimization of the growth conditions resulted in an almost two-fold increase in the expression levels in the fermentation supernatant of P. pastoris as compared to the starting conditions. We investigated the effect of methanol concentration, batch phase time, and batch to induction transition on NoV VLP-NY-ESO-1 production. The optimized process included a glycerol transition phase during the first 2 h of induction and a methanol concentration set point of 4 g L−1 during induction. Utilizing the bioprocess monitor to control the glycerol and methanol concentrations during induction resulted in a maximum NoV VP1-NY-ESO-1 yield of 0.85 g L−1.

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