P35: Temporal analysis of Pichia stipitis proteome using shotgun proteomics as a fermentation monitoring technique

Fermentation organisms can be better understood using shotgun proteomics as a monitoring technique. We demonstrated a shotgun proteomic method to monitor the temporal Pichia stipitis proteome from lag, log to stationary phases using xylose as the sole carbon source. For the first time, the global profile of P. stipitis proteome was observed with 958 proteins throughout the fermentation. Near complete enzymes involved in glycolysis and citrate cycle were identified, while key enzymes in pentose phosphate pathway (XYL1) was also observed. Respiration and fermentation coincided and were activate during cell growth under aerobic conditions. Beta glucosidase family 3, glycogen debranching enzyme and glycoside hydrolase family 17 were observed indicating activity against cellobiose or xylobiose. Approximately 20 differentially expressed proteins based on spectral counts were observed between each growth phase. Ribosomal proteins (L7A, L4/L1e, RS23, L22/L17, 40S S4 (S7), 60S large subunit) and cytochrome c oxidase were repressed in log phase. Phosphatidylinositol-specific phospholipase C, enolase I, phosphoglucomutase, aconitase, mitochondrial were induced from lag to log phase, indicating the increase activity in amino acid metabolism, secondary metabolism and macromolecule synthesis activity. However, cytochrome-c oxidase activity was decreased while the cells should be shifting into more oxidative phosphrylation in log phase. From log to stationary phase, ribosomal proteins (L4/L1e, L5, L10 , L19A) were repressed in stationary phase. 3-phosphoglycerate kinase was induced in stationary phase exhibiting alcohol catabolic process. The dynamic of P. stipitis proteome was observed and identified during fermentation in this temporal study.