A proteomic analysis of the Pichia pastoris secretome in methanol-induced cultures Journal Article
| Authors: | Huang, C. J.; Damasceno, L. M.; Anderson, K. A.; Zhang, S.; Old, L. J.; Batt, C. A. |
| Article Title: | A proteomic analysis of the Pichia pastoris secretome in methanol-induced cultures |
| Abstract: | The secreted proteome of Pichia pastoris X-33 was investigated in methanol-induced cultures with a goal to enhance the secretion and purification of recombinant proteins. In a fed-batch fermentation at 30 °C, more host proteins were found in greater concentrations compared to cultures grown at 25 °C. Protein samples collected directly from the culture media at 25 °C, as well as separated by two-dimensional (2D) gel, were subjected to ESI-MS/MS analysis. A total of 75 proteins were identified in the media from different conditions including pre- and post-methanol induction and in a strain overexpressing a recombinant schistosomiasis vaccine, Sm14-C62V. The identified proteins include native secreted proteins and some intracellular proteins, most of which have low isoelectric points (pI∈<∈6). 2D gel analyses further revealed important characteristics, such as abundance, degradation, and glycosylation of these identified proteins in this proteome. Cell wall-associated proteins involved in cell wall biogenesis, structure, and modification comprised the majority of the secreted proteins which have been identified. Intracellular proteins such as alcohol oxidase and superoxide dismutase were also found in the proteome, suggesting some degree of cell lysis. However, both protocols show that their concentrations are significantly lower than the native secreted proteins. This study identifies proteins secreted or released into the culture media in the methanol-induced fermentation cultures of P. pastoris X-33 and suggests potential biotechnology applications based on the discovery of this proteome. © 2011 Springer-Verlag. |
| Keywords: | nonhuman; mass spectrometry; proteins; proteome; protein degradation; oxygen; culture medium; protein; enzyme activity; biotechnology; proteomics; gels; cell wall; molecular sequence data; cell culture; protein purification; nucleotide sequence; recombinant protein; protein transport; protein secretion; cytolysis; yeast; electrophoresis, gel, two-dimensional; superoxide dismutase; vaccine; extracellular space; in-cell; gene expression regulation, fungal; secretome; culture media; secretion; two dimensional gel electrophoresis; recombination; methanol; fungal proteins; pichia pastoris; p. pastoris; fermentation; pichia; electrospray mass spectrometry; cell lysis; esi/ms/ms; methanol-induced culture; alcohol oxidase; biotechnology applications; cell walls; fed-batch fermentation; fermentation culture; gel analysis; host proteins; intracellular proteins; iso-electric points; protein samples; proteomes; proteomic analysis; secreted protein; super oxide dismutase; cell membrane protein; schistosoma vaccine; fed batch fermentation; fungus culture; protein glycosylation |
| Journal Title: | Applied Microbiology and Biotechnology |
| Volume: | 90 |
| Issue: | 1 |
| ISSN: | 0175-7598 |
| Publisher: | Springer |
| Date Published: | 2011-04-01 |
| Start Page: | 235 |
| End Page: | 247 |
| Language: | English |
| DOI: | 10.1007/s00253-011-3118-5 |
| PROVIDER: | scopus |
| PUBMED: | 21305280 |
| DOI/URL: | |
| Notes: | --- - "Export Date: 23 June 2011" - "CODEN: AMBID" - "Source: Scopus" |
