Beta-Propiolactone Inactivation of Coxsackievirus A16 Induces Structural Alteration and Surface Modification of Viral Capsids

Abstract

Beta-propiolactone (BPL) is an inactivating agent that is widely used in the vaccine industry. However, its effects on vaccine protein antigens and its mechanisms of action remain poorly understood. Here we present cryo-electron microscopy (cryo-EM) structures of BPL-treated coxsackievirus A16 (CVA16) mature virions and procapsids at resolutions of 3.9 Å and 6.5 Å, respectively. Notably, both particles were found to adopt an expanded conformation resembling the 135S-like uncoating intermediate, with characteristic features including an opened 2-fold channel, the externalization of the N terminus of VP1 capsid protein, and the absence of pocket factor. However, major neutralizing epitopes are very well preserved on these particles. Further biochemical analyses revealed that BPL treatment impairs the abilities of CVA16 particles to bind to the attachment receptor heparan sulfate and to a conformation-dependent monoclonal antibody in a BPL dose-dependent manner, indicating that BPL is able to modify surface-exposed amino acid residues. Taken together, our results demonstrate that BPL treatment may induce alteration of the overall structure and surface properties of a nonenveloped viral capsid, thus revealing a novel mode of action of BPL.IMPORTANCE Beta-propiolactone (BPL) is commonly used as an inactivating reagent to produce viral vaccines. It is recognized that BPL inactivates viral infectivity through modification of viral nucleic acids. However, its effect on viral proteins remains largely unknown. Here, we present high-resolution cryo-EM structures of BPLtreated coxsackievirus A16 (CVA16) mature virions and procapsids, which reveals an expanded overall conformation and characteristic features that are typical for the 135S-like uncoating intermediate. We further show that the BPL concentration affects the binding of inactivated CVA16 particles to their receptor/antibody. Thus, BPL treatment can alter the overall structure and surface properties of viral capsids, which may lead to antigenic and immunogenic variations. Our findings provide important information for future development of BPL-inactivated vaccines.KEYWORDS Beta-propiolactone, coxsackievirus A16, cryo-EM structure, structural alteration, surface modification E nterovirus 71 (EV71) and coxsackievirus A16 (CVA16) are the two major causative agents of hand, foot, and mouth disease (HFMD), which has been prevalent in young children in the Asia-Pacific region (1). According to a recent survey, EV71 and CVA16 were associated with 50.4% and 38.3% of the 266 laboratory-confirmed HFMD

A 3.0-Angstrom Resolution Cryo-Electron Microscopy Structure and Antigenic Sites of Coxsackievirus A6-Like Particles

“…The present study reveals that the CVA6 VLP consists of 60 protomers, each of which is composed of VP0 (the precursor of VP2 and VP4), VP1, and VP3 subunits. Similar findings have been reported for mature virions, procapsids, and VLPs of other enteroviruses (e.g., EV71 and CVA16) (23,25,27,40). However, the mechanisms underlying the assembly of protomers and particles of enteroviruses remain unclear.…”

Detection of chikungunya virus‐specific IgM on laser‐cut paper‐based device using pseudo‐particles as capture antigen

“…It is generally recognized that BPL inactivates viruses through cross‐linking of viral surface proteins. Consequently, BPL treatment could alter the overall structure of envelope viral proteins, which may modulate the affinity of antibodies for these proteins and thus decrease the sensitivity of the test …”

“…Consequently, BPL treatment could alter the overall structure of envelope viral proteins, which may modulate the affinity of antibodies for these proteins and thus decrease the sensitivity of the test. 60,61 To overcome these drawbacks, alternative antigens to the viral lysate were produced and tested for specific CHIKV IgM detection. 43 Among these antigens, VLPs and PPs, two types of synthetic viruses, were chosen because of their innocuous nature due to the absence of viral genome.…”

Advances in Antigenic Peptide-Based Vaccine and Neutralizing Antibodies against Viruses Causing Hand, Foot, and Mouth Disease

“…However, the large-scale growth and production of either of the viruses or the VLPs constituting the bivalent vaccine will not be cost effective. Moreover, chemical treatments have been shown to alter the antigenic surface of CV-A16 [32,51]. Considering these problems, it is better to produce a multi-epitope antigen or a chimeric antigen that can elicit neutralizing antibodies against both EV-A71 and CV-A16.…”