Surface Display of Rhizomucor Miehei Lipase in Pichia pastoris Using Sed1p as an Anchor Protein and Its Application

Abstract:

We constructed a Rhizomucor Miehei Lipase(RML)-Displaying yeast whole-cell biocatalyst and applied it to methylesters synthesized from triglyceride and methanol. RML was fused with the Sed1p cloned from Saccharomyces cerevisiae to constructed plasmid pPIC9K-Flag-RML-Sed1. The plasmid was linearized and transformed into Pichia pastoris GS115 and Pichia pastoris recombinant strain GS115/pPIC9K-Flag-RML-Sed1 was obtained. Cell-surface display of the RML via Sed1p was confirmed by flow Fluorescence micrograph and flow cytometer. After incubated at 28°C for 48 h the hydrolytic activity of the GS115/pPIC9K-Flag-RML-Sed1 reached a plateau, 169.6 U/g (Dry cell weight). In nonaqeous media, the yield of 82.36% methylesters was obtained from triglyceride and methanol after 72 h using lyophilized RML displaying yeast whole cells.