Abstract
Protein secretion in yeast is a complex process and its efficiency depends on a variety of parameters. We performed a comparative proteome analysis of a set of Schizosaccharomyces pombe strains producing the α-glucosidase maltase in increasing amounts to investigate the overall proteomic response of the cell to the burden of protein production along the various steps of protein production and secretion. Proteome analysis of these strains, utilizing an isobaric labeling/two dimensional LC-MALDI MS approach, revealed complex changes, from chaperones and secretory transport machinery to proteins controlling transcription and translation. We also found an unexpectedly high amount of changes in enzyme levels of the central carbon metabolism and a significant up-regulation of several amino acid biosyntheses. These amino acids were partially underrepresented in the cellular protein compared with the composition of the model protein. Additional feeding of these amino acids resulted in a 1.5-fold increase in protein secretion. Membrane fluidity was identified as a second bottleneck for high-level protein secretion and addition of fluconazole to the culture caused a significant decrease in ergosterol levels, whereas protein secretion could be further increased by a factor of 2.1. In summary, we show that high level protein secretion causes global changes of protein expression levels in the cell and that precursor availability and membrane composition limit protein secretion in this yeast. In this respect, comparative proteome analysis is a powerful tool to identify targets for an efficient increase of protein production and secretion in S. pombe Data are available via ProteomeXchange with identifiers PXD002693 and PXD003016.
Highlights
From the ‡Institute for Experimental Medicine, Div
We selected two of these targets, amino acid biosynthesis and membrane fluidity, which could be influenced via supplementation of the media with specific compounds in order to validate the efficacy of the proteome data for increasing protein secretion in S. pombe
Proteomic Response of the Central Metabolism to Elevated Levels of Maltase Secretion—Of the differentially abundant proteins identified in strain NW9 and NW10 compared with the wildtype, a large number were found to be related to central energy metabolism and anabolic pathways
Summary
From the ‡Institute for Experimental Medicine, Div. Systematic Proteome Research & Bioanalytics, Christian-Albrechts-Universitat zu Kiel, 24105 Kiel, Germany; §Biochemical Engineering Institute, Saarland University, 66123 Saarbrucken, Germany; ¶Roche Diagnostics GmbH, 68305 Mannheim, Germany; ʈPomBioTech GmbH, 66123 Saarbrucken, Germany; **School of Pharmaceutical Science and Technology, Tianjin University, Tianjin 300072, P.R. We selected two of these targets, amino acid biosynthesis and membrane fluidity, which could be influenced via supplementation of the media with specific compounds in order to validate the efficacy of the proteome data for increasing protein secretion in S. pombe.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
