Is Pichia pastoris a realistic platform for industrial production of recombinant human interferon gamma?

- Consistently high yield expression of hIFNγ in Pichia pastoris is not generally achievable.
- Further optimising of hIFNγ expression in P. pastoris did not result in viable commercial production potential.
- Lowering the minimal free energy of the mRNA did yield a 10-fold enhancement of expression of hIFNγ.
- Low abundance of hIFNγ mRNA in the recombinant P. pastoris, might be the reason for low expression.
- Results suggest that mammalian expression systems yield higher hIFNγ expression with suitable glycosylation patterns.

Human interferon gamma (hIFNγ) is an important cytokine in the innate and adaptive immune system, produced commercially in Escherichia coli. Efficient expression of hIFNγ has been reported once for Pichia pastoris (Wang et al., 2014) - a proven heterologous expression system. This study investigated hIFNγ expression in P. pastoris replicating the previous study and expanding by using four different strains (X33: wild type; GS115: HIS−Mut+; KM71H: Arg+, Mut− and CBS7435: MutS) and three different vectors (pPICZαA, pPIC9 and pPpT4αS). In addition, the native sequence (NS) and two codon-optimised sequences (COS1 and COS2) for P. pastoris were used. Methanol induction yielded no expression/secretion of hIFNγ in X33, highest levels were recorded for CBS7435: MutS (∼16 μg. L−1). mRNA copy number calculations acquired from RT-qPCR for GS115-pPIC9-COS1 proved low abundance of mRNA. A 10-fold increase in expression of hIFNγ was achieved by lowering the minimal free energy of the mRNA and 100-fold by MutS phenotypes, substantially lower than reported by Wang et al. (2014). We conclude that commercial production of low cost, eukaryotic recombinant hIFNγ is not an economically viable in P. pastoris. Further research is required to unravel the cause of low expression in P. pastoris to achieve economic viability.