Effect of Fc Fusion on Folding and Immunogenicity of Middle East Respiratory Syndrome Coronavirus Spike Protein

Abstract

Middle East respiratory syndrome coronavirus (MERS-CoV) induces severe respiratory impairment with a reported mortality rate of ~36% in humans. The absence of clinically available MERS-CoV vaccines and treatments to date has resulted in uncontrolled incidence and propagation of the virus. In vaccine design, fusion with the IgG Fc domain is reported to increase the immunogenicity of various vaccine antigens. However, limited reports have documented the potential negative effects of Fc fusion on vaccine antigens. To determine whether Fc fusion affects the immunogenicity of MERS-CoV antigen, we constructed a Fcassociated MERS-CoV spike protein (eS770-Fc, 110 kDa), whereby human IgG4 Fc domain was fused to MERS-CoV spike protein (eS770) via a Gly/Pro linker using baculovirus as the expression system. For comparative analyses, two eS770 proteins lacking the IgG4 Fc domain were generated using the IdeS protease (eS770-ΔFc) or His tag attachment (eS770-His) and the immunogenicity of the above constructs were examined following intramuscular immunization in mice. Contrary to expectations, non-Fc spike proteins (eS770-ΔFc, eS770-His; 90 kDa) showed higher immunogenicity than the Fc fusion protein (eS770-Fc). Moreover, unlike non-Fc spike proteins, eS770-Fc immunization did not elicit neutralizing antibodies against MERS-CoV. The lower immunogenicity of Fc-fused eS770 was related to alterations in the structural conformation of the spike protein. Taken together, our results indicate that IgG Fc fusion reduces the immunogenicity of eS770 by interfering with the proper folding structure.

Rapid, high-yield production of full-length SARS-CoV-2 spike ectodomain by transient gene expression in CHO cells

“…It is also possible to express the full-length spike, including transmembrane and Cterminal domains, which can be purified following membrane solubilisation of expressing cells [9,10]. Finally, expression of the full-length soluble forms of the coronavirus spike ECDs has also been reported in HEK293 and insect cells [6,[11][12][13][14][15]. Importantly, however, in cases where this data is reported, yields were extremely low, ranging from 0.5-1.5 mg per litre of culture media (for expression of constructs containing the full-length ECD) [9,14].…”

Codelivery of improved immune complex and virus-like particle vaccines containing Zika virus envelope domain III synergistically enhances immunogenicity

“…Important epitopes may be obscured or improperly folded depending on interactions between the antigen and the RICs or VLPs. Fusion of the Middle East respiratory syndrome coronavirus spike protein to IgG Fc interfered with protein folding, leading to reduced immunogenicity [56]. Fig.…”

“…Based on our observations, RIC and VLP combinations have been shown to enhance neutralizing antibody titers against ZE3 (described here), HPV L2 [24], and influenza M2e (data to be presented elsewhere), indicating that the enhancement is not antigen-specific. RICs activate complement C1q and elicit Fc-mediated effector functions [26], while HBc VLPs contain strong T-cell epitopes, potently activate macrophages, and may stimulate TLRs due to encapsidated nucleic acid [56]. Therefore, providing both RICs and VLPs simultaneously may stimulate more arms of the immune system, producing a multiplicative rather than additive effect.…”

Human endogenous retrovirus-enveloped baculoviral DNA vaccines against MERS-CoV and SARS-CoV2

“…Neutralization titers were defined as the reciprocal of the highest serum dilution at which luciferase activity was reduced by at least 50%. Neutralization assays were performed as previously reported 31 .…”