An Efficient Genome-Wide Fusion Partner Screening System for Secretion of Recombinant Proteins in Yeast

Overview

Journal Sci Rep

Specialty Science

Date 2015 Jul 22

PMID 26195161

Citations 21

Authors

Jung-Hoon Bae

Bong Hyun Sung

Hyun-Jin Kim

Soon-Ho Park

Kwang-Mook Lim

Mi-Jin Kim

Cho-Ryong Lee

Jung-Hoon Sohn

Affiliations

Soon will be listed here.

Abstract

To produce rarely secreted recombinant proteins in the yeast Saccharomyces cerevisiae, we developed a novel genome-wide optimal translational fusion partner (TFP) screening system that involves recruitment of an optimal secretion signal and fusion partner. A TFP library was constructed from a genomic and truncated cDNA library by using the invertase-based signal sequence trap technique. The efficiency of the system was demonstrated using two rarely secreted proteins, human interleukin (hIL)-2 and hIL-32. Optimal TFPs for secretion of hIL-2 and hIL-32 were easily selected, yielding secretion of these proteins up to hundreds of mg/L. Moreover, numerous uncovered yeast secretion signals and fusion partners were identified, leading to efficient secretion of various recombinant proteins. Selected TFPs were found to be useful for the hypersecretion of other recombinant proteins at yields of up to several g/L. This screening technique could provide new methods for the production of various types of difficult-to-express proteins.

Citing Articles

Engineering for efficient production of recombinant proteins.

Yang S, Song L, Wang J, Zhao J, Tang H, Bao X Eng Microbiol. 2024; 4(1):100122.

PMID: 39628786 PMC: 11611019. DOI: 10.1016/j.engmic.2023.100122.

A yeast surface display platform for screening of non-enzymatic protein secretion in Kluyveromyces lactis.

An J, Shang N, Liu W, Niu Y, Liang Q, Jiang J Appl Microbiol Biotechnol. 2024; 108(1):503.

PMID: 39500795 PMC: 11538148. DOI: 10.1007/s00253-024-13342-9.

oral immunization in mice using multi-antigen of the African swine fever virus elicits a robust immune response.

Gao S, Zuo W, Kang C, Zou Z, Zhang K, Qiu J Front Immunol. 2024; 15:1373656.

PMID: 38742108 PMC: 11089227. DOI: 10.3389/fimmu.2024.1373656.

Tailoring in fungi for next generation cellulase production with special reference to CRISPR/CAS system.

Mondal S, Halder S, Mondal K Syst Microbiol Biomanuf. 2024; 2(1):113-129.

PMID: 38624901 PMC: 8319711. DOI: 10.1007/s43393-021-00045-9.

A Yeast Modular Cloning (MoClo) Toolkit Expansion for Optimization of Heterologous Protein Secretion and Surface Display in .

ORiordan N, Juric V, ONeill S, Roche A, Young P ACS Synth Biol. 2024; 13(4):1246-1258.

PMID: 38483353 PMC: 11036508. DOI: 10.1021/acssynbio.3c00743.

References

Rakestraw J, Sazinsky S, Piatesi A, Antipov E, Wittrup K . Directed evolution of a secretory leader for the improved expression of heterologous proteins and full-length antibodies in Saccharomyces cerevisiae. Biotechnol Bioeng. 2009; 103(6):1192-201. PMC: 2847895. DOI: 10.1002/bit.22338. View

Mormeneo M, Andres I, Bofill C, Diaz P, Zueco J . Efficient secretion of Bacillus subtilis lipase A in Saccharomyces cerevisiae by translational fusion to the Pir4 cell wall protein. Appl Microbiol Biotechnol. 2008; 80(3):437-45. DOI: 10.1007/s00253-008-1549-4. View

Smith J, Tang B, Robinson A . Protein disulfide isomerase, but not binding protein, overexpression enhances secretion of a non-disulfide-bonded protein in yeast. Biotechnol Bioeng. 2004; 85(3):340-50. DOI: 10.1002/bit.10853. View

Granovskaia M, Jensen L, Ritchie M, Toedling J, Ning Y, Bork P . High-resolution transcription atlas of the mitotic cell cycle in budding yeast. Genome Biol. 2010; 11(3):R24. PMC: 2864564. DOI: 10.1186/gb-2010-11-3-r24. View

Kushnirov V . Rapid and reliable protein extraction from yeast. Yeast. 2000; 16(9):857-60. DOI: 10.1002/1097-0061(20000630)16:9<857::AID-YEA561>3.0.CO;2-B. View

Bleve G, Lezzi C, Spagnolo S, Rampino P, Perrotta C, Mita G . Construction of a laccase chimerical gene: recombinant protein characterization and gene expression via yeast surface display. Appl Biochem Biotechnol. 2014; 172(6):2916-31. DOI: 10.1007/s12010-014-0734-4. View

Kim S, Han S, Azam T, Yoon D, Dinarello C . Interleukin-32: a cytokine and inducer of TNFalpha. Immunity. 2005; 22(1):131-42. DOI: 10.1016/j.immuni.2004.12.003. View

Ahn J, Park K, Lee H, Son Y, Choi E . GAL promoter-driven heterologous gene expression in Saccharomyces cerevisiae Δ strain at anaerobic alcoholic fermentation. FEMS Yeast Res. 2012; 13(1):140-2. DOI: 10.1111/1567-1364.12009. View

Mizuno K, Nakamura T, Ohshima T, Tanaka S, Matsuo H . Yeast KEX2 genes encodes an endopeptidase homologous to subtilisin-like serine proteases. Biochem Biophys Res Commun. 1988; 156(1):246-54. DOI: 10.1016/s0006-291x(88)80832-5. View

10.

Werten M, van den Bosch T, Wind R, Mooibroek H, de Wolf F . High-yield secretion of recombinant gelatins by Pichia pastoris. Yeast. 1999; 15(11):1087-96. DOI: 10.1002/(SICI)1097-0061(199908)15:11<1087::AID-YEA436>3.0.CO;2-F. View

11.

Sievi E, Hanninen A, Salo H, Kumar V, Makarow M . Validation of the Hsp150 polypeptide carrier and HSP150 promoter in expression of rat alpha2,3-sialyltransferase in yeasts. Biotechnol Prog. 2003; 19(4):1368-71. DOI: 10.1021/bp034035p. View

12.

Kjeldsen T, Pettersson A, Hach M, Diers I, Havelund S, Hansen P . Synthetic leaders with potential BiP binding mediate high-yield secretion of correctly folded insulin precursors from Saccharomyces cerevisiae. Protein Expr Purif. 1997; 9(3):331-6. DOI: 10.1006/prep.1996.0695. View

13.

Gietz R, Schiestl R, Willems A, Woods R . Studies on the transformation of intact yeast cells by the LiAc/SS-DNA/PEG procedure. Yeast. 1995; 11(4):355-60. DOI: 10.1002/yea.320110408. View

14.

Mormeneo M, Pastor F, Zueco J . Efficient expression of a Paenibacillus barcinonensis endoglucanase in Saccharomyces cerevisiae. J Ind Microbiol Biotechnol. 2011; 39(1):115-23. DOI: 10.1007/s10295-011-1006-8. View

15.

Anzur Lasnik M, Nemec M, Romih R, Schara M . An EPR study of the secretion of G-CSF heterologous protein from Pichia pastoris. Biotechnol Bioeng. 2003; 81(7):768-74. DOI: 10.1002/bit.10527. View

16.

Choi E, Sohn J, Rhee S . Optimization of the expression system using galactose-inducible promoter for the production of anticoagulant hirudin in Saccharomyces cerevisiae. Appl Microbiol Biotechnol. 1994; 42(4):587-94. DOI: 10.1007/BF00173925. View

17.

Han K, Kim S, Choi S, Seong B . N-Glycosylation of secretion enhancer peptide as influencing factor for the secretion of target proteins from Saccharomyces cerevisiae. Biochem Biophys Res Commun. 2005; 337(2):557-62. DOI: 10.1016/j.bbrc.2005.09.089. View

18.

Salo H, Sievi E, Suntio T, Mecklin M, Mattila P, Renkonen R . Co-expression of two mammalian glycosyltransferases in the yeast cell wall allows synthesis of sLex. FEMS Yeast Res. 2005; 5(4-5):341-50. DOI: 10.1016/j.femsyr.2004.11.007. View

19.

Simonen M, Jamsa E, Makarow M . The role of the carrier protein and disulfide formation in the folding of beta-lactamase fusion proteins in the endoplasmic reticulum of yeast. J Biol Chem. 1994; 269(19):13887-92. View

20.

Andres I, Rodriguez-Diaz J, Buesa J, Zueco J . Yeast expression of the VP8* fragment of the rotavirus spike protein and its use as immunogen in mice. Biotechnol Bioeng. 2005; 93(1):89-98. DOI: 10.1002/bit.20696. View