Characterization of the Bgs13 protein’s role in the | 40287
Journal of Proteomics & Bioinformatics

Journal of Proteomics & Bioinformatics
Open Access

ISSN: 0974-276X

+44 1223 790975

Characterization of the Bgs13 protein’s role in the super-secretion of recombinant peptides in the yeast Pichia pastoris

3rd International Conference on Antibodies, Bio Therapeutics & B2B & Genetic and Protein Engineering

November 08-09, 2017 | Las Vegas, USA

Geoff P Lin-Cereghino, Joan Lin-Cereghino, Anita D Jivan, Maria N Vo, Michelle M Fong and Douglas D Risser

University of the Pacific, USA

Posters & Accepted Abstracts: J Proteomics Bioinform

Abstract :

Statement of the Problem: The yeast Pichia pastoris is a popular host for expressing and exporting recombinant proteins, such as human insulin and a hepatitis B vaccine protein, out of its cell. Secreted proteins are easier to purify and therefore are more useful than non-secreted proteins. However, Pichia pastoris has been known to secrete certain proteins efficiently while struggling to secrete others. Approach & Methodology: Our lab has created a strain with a mutated Bgs13 gene that is a super-secreter of multiple recombinant peptides. To understand why the Bgs13 strain displays enhanced secretion, cell wall assays were first performed using Congo red and Calcoflour white to determine if super-secretion is a result of defective cell walls. In addition, Bgs13 appears to be a homolog of the Saccharomyces cerevisiae protein kinase C (PKC1). Thus, we tested if super-secretion in our Bgs13 strain is a result of elevated or decreased protein kinase C activity compared to the wild type parent. Lastly, the localization of wild type Bgs13 and mutant Bgs13 proteins was compared by fusing each protein to EGFP and examining them with fluorescence microscopy analysis. Results & Significance: The mutant Bgs13 strain had a cell wall with apparent structural defects. Not only did the mutant Bgs13 protein have lower protein kinase C activity, but it also was localized to different parts of the P. pastoris cell compared to the wild type Bgs13 protein. By characterizing mutant and wild type Bgs13 proteins, the results will help us create strains with optimized secretion of many different recombinant proteins.

Biography :

Geoff Lin-Cereghino supervise a lab of undergraduates and Masters students at the University of the Pacific in Stockton, California. His work focusses on investigating the secretory mechanism of the yeast Pichia pastoris in order to improve the production of recombinant proteins in this host. Most recently, his research has concentrated on characterizing a group of super-secreting mutant strains isolated in his lab as well as optimizing the secretion efficiency of the α-mating factor prepro-peptide, which is the most commonly used secretion signal for heterologous protein expression in P.pastoris. He have been recipients of NIH AREA and NIH RUI grants.