High-level heterologous expression of the human transmembrane sterol Δ8,Δ7-isomerase in Pichia pastoris.

Recombinant expression of human membrane proteins in large quantities remains a major challenge. Expression host is an important variable to screen for high-level production of membrane proteins. Using the green fluorescent protein (GFP) as a reporter, we screened the expression of a human multi-pass membrane protein called sterol Δ8-Δ7 isomerase in three different hosts: Escherichia coli, Saccharomyces cerevisiae, and Pichia pastoris. The expression of the His-tagged isomerase was exceptionally high in P. pastoris, reaching ~200 mg L-1 in standard flasks, and ~1,000 mg L-1 in condensed culture that mimics fermentation. The heterogeneously expressed isomerase could be extracted fully with dodecyl maltoside, and the solubilized protein in the form of GFP fusion showed a sharp and symmetric peak on fluorescence-detection size exclusion chromatography. Our work provides a useful source for the purification of the recombinant isomerase.