Tyo, Keith E. J.3; Liu, Zihe3; Magnusson, Ylva3; Petranovic, Dina4; Nielsen, Jens1
1 Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark2 Fungal Cell Factories, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark3 Chalmers University of Technology4 Department of Systems Biology, Technical University of Denmark
Protein titers, a key bioprocessing metric, depend both on the synthesis of protein and the degradation of protein. Secreted recombinant protein production in Saccharomyces cerevisiae is an attractive platform as minimal media can be used for cultivation, thus reducing fermentation costs and simplifying downstream purification, compared to other systems that require complex media. As such, engineering S. cerevisiae to improve titers has been then the subject of significant attention, but the majority of previous efforts have been focused on improving protein synthesis. Here, we characterize the protein uptake and degradation pathways of S. cerevisiae to better understand its impact on protein secretion titers. We do find that S. cerevisiae can consume significant (in the range of 1 g/L/day) quantities of whole proteins. Characterizing the physiological state and combining metabolomics and transcriptomics, we identify metabolic and regulatory markers that are consistent with uptake of whole proteins by endocytosis, followed by intracellular degradation and catabolism of substituent amino acids. Uptake and degradation of recombinant protein products may be common in S. cerevisiae protein secretion systems, and the current data should help formulate strategies to mitigate product loss.
Applied Microbiology and Biotechnology, 2014, Vol 98, Issue 16, p. 7149-7159
endocytosis; protein degradation pathway; protein uptake; Fungi Plantae (Fungi, Microorganisms, Nonvascular Plants, Plants) - Ascomycetes  Saccharomyces cerevisiae species yeast common strain-CEN.PK 113-7D; amino acid catabolism intracellular degradation; metabolic marker; recombinant protein; regulatory marker; 10060, Biochemistry studies - General; 10064, Biochemistry studies - Proteins, peptides and amino acids; 39008, Food microbiology - General and miscellaneous; 51522, Plant physiology - Chemical constituents; Biochemistry and Molecular Biophysics; Bioprocess Engineering; BIOTECHNOLOGY; SACCHAROMYCES-CEREVISIAE; METABOLOMICS; EXTRACTION; TOPOLOGY; SOFTWARE; RATES; Endocytosis; Saccharomyces cerevisiae; Protein degradation; Protein production; Secretion; PROTEIN synthesis; HASH(0x34dfcd0)