Enhanced cell-surface display and secretory production of cellulolytic enzymes with Saccharomyces cerevisiae Sed1 signal peptide

• Published in: Biotechnology and bioengineering Vol. 113; no. 11; pp. 2358 - 2366
• Main Authors: Inokuma, Kentaro, Bamba, Takahiro, Ishii, Jun, Ito, Yoichiro, Hasunuma, Tomohisa, Kondo, Akihiko
• Format: Journal Article
• Language: English
• Published: United States Blackwell Publishing Ltd 01.11.2016; Wiley Subscription Services, Inc
• Subjects: Anchoring; Aspergillus aculeatus; Baking yeast; Bioengineering; Biofuels; Cassettes; Cell Membrane - metabolism; Cell surface; cell surface display; Cell walls; Cellular biology; Cellulase - secretion; Cellulolytic enzymes; endo-glucanase; Endoglucanase; Enzymes; Ethanol; EXG1 protein; Fungi; Gene sequencing; Genes; Genetic Enhancement - methods; Genomes; Glucoamylase; Glucosidase; Glycosylphosphatidylinositol; Hypocrea jecorina; Lignocellulose; Localization; Mating; Membrane Glycoproteins - genetics; Membrane Glycoproteins - metabolism; Peptides; Pichia pastoris; Protein Engineering - methods; Protein Transport - genetics; Proteins; Recombinant; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Rhizopus oryzae; Saccharomyces cerevisiae; Saccharomyces cerevisiae - physiology; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Secretion; secretion signal sequence; Surface chemistry; Utilization; Yeast; β-glucosidase; Pichia pastoris; β-glucosidase; endo-glucanase; Saccharomyces cerevisiae; secretion signal sequence; cell surface display
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.26008

ABSTRACT Recombinant yeast strains displaying aheterologous cellulolytic enzymes on their cell surfaces using a glycosylphosphatidylinositol (GPI) anchoring system are a promising strategy for bioethanol production from lignocellulosic materials. A crucial step for cell wall localization of the enzymes is the intracellular transport of proteins in yeast cells. Therefore, the addition of a highly efficient secretion signal sequence is important to increase the amount of the enzymes on the yeast cell surface. In this study, we demonstrated the effectiveness of a novel signal peptide (SP) sequence derived from the Saccharomyces cerevisiae SED1 gene for cell‐surface display and secretory production of cellulolytic enzymes. Gene cassettes with SP sequences derived from S. cerevisiae SED1 (SED1SP), Rhizopus oryzae glucoamylase (GLUASP), and S. cerevisiae α‐mating pheromone (MFα1SP) were constructed for cell‐surface display of Aspergillus aculeatus β‐glucosidase (BGL1) and Trichoderma reesei endoglucanase II (EGII). These gene cassettes were integrated into the S. cerevisiae genome. The recombinant strains with the SED1SP showed higher cell‐surface BGL and EG activities than those with the conventional SP sequences (GLUASP and MFα1SP). The novel SP sequence also improved the secretory production of BGL and EG in S. cerevisiae. The extracellular BGL activity of the recombinant strains with the SED1SP was 1.3‐ and 1.9‐fold higher than the GLUASP and MFα1SP strains, respectively. Moreover, the utilization of SED1SP successfully enhanced the secretory production of BGL in Pichia pastoris. The utilization of the novel SP sequence is a promising option for highly efficient cell‐surface display and secretory production of heterologous proteins in various yeast species. Biotechnol. Bioeng. 2016;113: 2358–2366. © 2016 Wiley Periodicals, Inc. The effectiveness of a novel signal peptide (SP) sequence derived from the Saccharomyces cerevisiae SED1 gene (SED1SP) for cell‐surface display and secretory production of heterologous enzymes is described. High secretion efficiency of the novel SP sequence was applicable for both cell‐surface display and secretory production of cellulolytic enzymes (BGL and EG) in S. cerevisiae. In addition, the utilization of SED1SP successfully enhanced the secretory production of BGL in Pichia pastoris.