Impact of NH4+ nitrogen source on the production of Rhizopus oryzae lipase in Pichia pastoris

Abstract Background Pichia pastoris is a highly successful system for heterologous expression. During the induction stage, the ammonium ion released into the fermentation broth has a deep impact on cell growth and protein expression. The impact of NH 4 + concentration on the expression of the Rhizopus oryzae lipase proAROL in P. pastoris was investigated. Results The lipase activity under the optimum NH 4 + concentration of 440 mmol/L reached 12,019 U/mL. Increased concentrations of NH 4 + in the broth prevented the protease production, resulting in higher specific lipase activity in the supernatant. Furthermore, analysis of carbon metabolism and energy regeneration pattern revealed that under the definite NH 4 + concentrations more carbon source (methanol) was consumed with surged AOX activity and then the higher energy and amino acid precursors demand for recombinant protein synthesis is compensated for by the TCA cycle. Conclusions In this study, the R. oryzae lipase activity reaches the highest level ever reported under optimized NH 4 + concentration and the analysis of the carbon metabolism provides useful information for future optimization of protein production by P. pastoris in a molecular level.