Alanine Substitution Inactivates Cross-Reacting Epitopes in Dengue Virus Recombinant Envelope Proteins.

Viviana C Zomosa-Signoret,Román Vidaltamayo,Ana M Rivas-Estilla,Edgar Galaviz-Aguilar,Ana E Estrada-Rodríguez,Karina R Morales-González,M Cristina Devèze-García

doi:10.3390/v12020208

Viviana C Zomosa-Signoret, Román Vidaltamayo + Show 5 more

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https://doi.org/10.3390/v12020208

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Abstract

The expansion of the habitat of mosquitoes belonging to the Aedes genus puts nearly half of the world’s population at risk of contracting dengue fever, and a significant fraction will develop its serious hemorrhagic complication, which can be fatal if not diagnosed properly and treated in a timely fashion. Although several diagnostic methods have been approved for dengue diagnostics, their applicability is limited in rural areas of developing countries by sample preparation costs and methodological requirements, as well as cross-reactivity among the different serotypes of the Dengue virus and other flavivirus, such as the Zika virus. For these reasons, it is necessary to generate more specific antigens to improve serological methods that could be cheaper and used in field operations. Here, we describe a strategy for the inactivation of cross-reacting epitopes on the surface of the Dengue virus envelope protein through the synthetic generation of recombinant peptide sequences, where key amino acid residues from Dengue virus serotype 1 (DENV-1) and 2 (DENV-2) are substituted by alanine residues. The proteins thus generated are recognized by 88% of sera from Dengue NS1+ patients and show improved serotype specificity because they do not react with the antibodies present in seroconverted, PCR-serotyped DEN-4 infected patients.

Highlights

Dengue and other mosquito-borne viral diseases threaten heavily populated areas around the world, and the expansion of the habitat of their vector and an increase in human mobility lead to the introduction of novel infectious agents, which pose new challenges to national health services, as the rapid spread of Chikungunya and Zika viruses in the Americas has demonstrated [1,2].Four serotypes of Dengue viruses (DENV1 through DENV4) have been identified
Those substitutions that lead to the alteration of the solvent accessible structure of the cross-reacting epitopes, with minor perturbations to the overall 3D structure of the protein model, were selected for the generation of mutant versions of the E proteins through synthetic DNA sequence generation and expression in an E. coli heterologous system
Most of the mosquito-airborne viruses, and their vectors, are endemic to regions of the world where economic development prevents the widespread use of molecular diagnostic methods that allow for the identification and serotypification of the infecting virus

Summary

Introduction

Dengue and other mosquito-borne viral diseases threaten heavily populated areas around the world, and the expansion of the habitat of their vector and an increase in human mobility lead to the introduction of novel infectious agents, which pose new challenges to national health services, as the rapid spread of Chikungunya and Zika viruses in the Americas has demonstrated [1,2].Four serotypes of Dengue viruses (DENV1 through DENV4) have been identified. In naturally occurring infections, acquired immunity against one serotype rarely protects against a subsequent secondary infection by any of the other serotypes [3,4,5], but the presence of circulating, non-neutralizing antibodies against any of the serotypes increases the risk of severe hemorrhagic fever/shock syndrome in patients infected by a different serotype in a secondary or concurrent exposure [4,5,6]. The envelope E protein of the DENV virion is a major antigen determinant of its serotypes and serves as a target for most of the antibodies generated by patients in naturally occurring infections [5,7]. Environmental acidification upon cell internalization into the endocytic pathway leads to a conformational change of the E proteins that trimerize at low pH, which allows for the insertion of the E protein into the membrane of the endolysosomes and the escape of the genetic RNA material of the virion into the cytosol [8,9,11,12]

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