title: Enhancing antibody folding and secretion by tailoring the Saccharomyces cerevisiae endoplasmic reticulum.
abstract: BACKGROUND
  The yeast Saccharomyces cerevisiae provides intriguing possibilities for synthetic biology and bioprocess applications, but its use is still constrained by cellular characteristics that limit the product yields. Considering the production of advanced biopharmaceuticals, a major hindrance lies in the yeast endoplasmic reticulum (ER), as it is not equipped for efficient and large scale folding of complex proteins, such as human antibodies.
  
  
  RESULTS
  Following the example of professional secretory cells, we show that inducing an ER expansion in yeast by deleting the lipid-regulator gene OPI1 can improve the secretion capacity of full-length antibodies up to fourfold. Based on wild-type and ER-enlarged yeast strains, we conducted a screening of a folding factor overexpression library to identify proteins and their expression levels that enhance the secretion of antibodies. Out of six genes tested, addition of the peptidyl-prolyl isomerase CPR5 provided the most beneficial effect on specific product yield while PDI1, ERO1, KAR2, LHS1 and SIL1 had a mild or even negative effect to antibody secretion efficiency. Combining genes for ER enhancement did not induce any significant additional effect compared to addition of just one element. By combining the Δopi1 strain, with the enlarged ER, with CPR5 overexpression, we were able to boost the specific antibody product yield by a factor of 10 relative to the non-engineered strain.
  
  
  CONCLUSIONS
  Engineering protein folding in vivo is a major task for biopharmaceuticals production in yeast and needs to be optimized at several levels. By rational strain design and high-throughput screening applications we were able to increase the specific secreted antibody yields of S. cerevisiae up to 10-fold, providing a promising strain for further process optimization and platform development for antibody production.
authors.affiliation: Department of Biotechnology and Chemical Technology, Aalto University, Kemistintie 1, 02150, Espoo, Finland.
authors.forename: Jorg C
authors.initials: JC
authors.lastname: de Ruijter
authors.affiliation: Department of Biotechnology and Chemical Technology, Aalto University, Kemistintie 1, 02150, Espoo, Finland.
authors.forename: Essi V
authors.initials: EV
authors.lastname: Koskela
authors.affiliation: Department of Biotechnology and Chemical Technology, Aalto University, Kemistintie 1, 02150, Espoo, Finland. alexander.frey@aalto.fi.
authors.forename: Alexander D
authors.initials: AD
authors.lastname: Frey
mesh_terms: D000906:Antibodies / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D019696:Peptidylprolyl Isomerase / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D017510:Protein Folding; D012441:Saccharomyces cerevisiae / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013696:Temperature
chemical_list: D000906:Antibodies; D029701:Saccharomyces cerevisiae Proteins; D019696:Peptidylprolyl Isomerase
keywords: Antibody; Chaperones; Endoplasmic reticulum; Heterologous protein production; Protein folding