BIOMEDICAL (APPLIED/EXPLORATORY)

Lifesensors, Inc., Malvern PA

Investigators

Abstract

Most human proteins are glycosylated. Increasing demand for recombinant glycosylated proteins of therapeutic and diagnostic importance has focused research on techniques for improving protein expression and controlling post-translational processing. We propose to use SUMO-fusion vector to improve protein expression and protein secretion and engineered humanized P.pastoris strain to control post-translational glycosylation. In Phase I, we will demonstrate that our novel system (i) is superior in increasing protein expression and secretion, facilitating protein purification and generating desired N-terminal amino acid, (ii) produce proteins with more than 90% homogeneity, (iii) produce proteins with mammalian-like N-glycan complex structures. As a proof-oF-principle we wi ll express and purify 10 cancer-related glycosylated proteins. Purified glycoproteins will be analyzed using DSA-FACE or mass spec to demonstrate mammalian-like post~translatjonal glycosylation. In Phase II we will use our developed system and establislled growth conditions to express, purify and characterize an additional 100 cancer-related glycoproteins. Our novel protein expression system not only improves protein expression/purification and controls post-translational glycosylation, but also reduces the time and cost of R&D, accelerating process from genomics to proteomics to therapeutic proteins.