Expression cassette and plasmid construction for Yeast Surface Display in Saccharomyces cerevisiae

Abstract

Objectives. Develop a Cell Surface Display system in S. cerevisiae, based on the construction of an expression cassette for pYES2 plasmid.Results. The construction of an expression cassette containing the α-factor signal peptide and the Cterminal portion of the α-agglutinin protein was made and its sequence inserted into a plasmid named pYES2/gDαAgglutinin, allowing cell surface display of bovine herpesvirus type 5 (BoHV-5) glycoprotein D (gD) on S. cerevisiae BY4741 strain. Recombinant protein expression was con rmed by dot blot, and indirect immuno uorescence using monoclonal anti-histidine antibodies and polyclonal antibodies from mice experimentally vaccinated with a recombinant gD.Conclusions. These results demonstrate that the approach and plasmid used represent not only an effective system for immobilizing proteins on the yeast cell surface, as well as a platform for immunobiologicals development.

Platforms for Production of Protein-Based Vaccines: From Classical to Next-Generation Strategies

“…Antigens from virus, protozoa, bacteria, nematode and tick expressed in different species of yeast, including P. pastoris, have been extensively reviewed by Kumar and colleagues [241]. Recently, it has been reported that Zika virus envelope protein (80E) expressed in P. pastoris is able to elicit potent ZIKV-neutralizing antibodies in mice conferring significant protection in vivo [242]. SARS-CoV-2 recombinant receptor-binding domain candidate vaccine expressed in P. pastoris has also been reported to be able to stimulate virus neutralizing antibodies and T-cell immunity in mice [242].…”

“…New approaches in recombinant yeast vaccine technology include the use of inactivated whole yeast cells [243] or antigen display at the yeast cell surface [242] as vaccination strategy. These technologies are of great interest for the development of oral vaccines at low-cost because no purification is needed.…”

“…Moreover, this delivery system could also enhance immunogenicity of vaccines due to the adjuvant activity of β-glucans on the yeast cell wall that have demonstrated immunomodulatory and adjuvant effects through the binding of innate pathogen receptors on macrophages, DC and neutrophils [245]. Some preclinical studies based on orally administrated S. cerevisiae displaying antigens on its surface for different infectious agents, such as influenza, showed that this delivery system is able to induce both humoral and cellular responses in mice [242]. Current developments include the expression of the spike protein of SARS-CoV-2 on the surface of S. cerevisiae as vaccine candidate, showing promising results in mice [246].…”