Journal of Virology & Antiviral ResearchISSN: 2324-8955

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Research Article, J Virol Antivir Res Vol: 4 Issue: 3

Occurrence of Escape Favoring Mutations in The Targets of 2F5 and 4E10 Antibodies of HIVgp41 : A Metadata Analysis

Christdas J and Shakila H*
Department ofMolecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Madurai, Tamilnadu, India
Corresponding author : H Shakila
Department of Molecular Microbiology, School of Biotechnology, Madurai Kamaraj University, Madurai, Tamilnadu, India,
Tel: 0452 - 2458723
E-mail: [email protected]
Received: July 17, 2015 Accepted: August 27, 2015 Published: September 02, 2015
Citation: Christdas J, Shakila H (2015) Occurrence of Escape Favoring Mutations in The Targets of 2f5 and 4e10 Antibodies of HIV gp41 : A Metadata Analysis. J Virol Antivir Res 4:3. doi:10.4172/2324-8955.1000141

Abstract

Objective

The preventive vaccine for HIV has been an unachieved goal and the increasing prevalence burdens the field of medicine for the last three decades. The challenge is due to the substantial overall sequence variability that is found across the genome of HIV- 1. Here a retrospection has been done to investigate the degree of conservation in the targets of the 2F5 and 4E10 neutralizing antibodies. The 6 and 7 amino acid long epitopes of the neutralizing antibodies are embedded in the MPER (Membrane Proximal External Region) of the HIV-1 glycoprotein 41.

Methods

HIV-1 gp 41 nucleotide sequences reported from different parts of the world were collected as fasta sequences on a random basis. The sequences were aligned and investigated for mutations found in the epitopes targeted by the 2F5 and 4E10 neutralizing antibodies.

Results

Substitutions and deletions almost in all the residues were found. The mutations that were found in each of the residue can critically affect the affinity of the neutralizing antibodies. The occurrence of these mutations in the critical residues in combination with the neighboring residues may offer a survival advantage for the virus to escape from these circulating neutralizing antibodies. This invariant nature which is substantiated earlier has been diverging over the years.

Conclusion

The mutations can crucially alter the dynamicity and the reactivity of the 2F5 and 4E10 antibodies directed to the putative vaccine targets. This confers the fact that these targets may be less appropriate a candidate for a vaccine. The infidel reverse transcriptase that copies these ‘selfish’ survivable mutants is characteristically favoring the virus to make progeny which can escape the surveillance by these neutralizing antibodies. Hence, due to the observed trend in the lack of conservation, it becomes necessary to search for better candidates for a stronger immune response that promises success in vaccine strategies.

Keywords: Neutralizing antibodies; HIV epitopes; Gp41 mutations; Vaccine targets; Bioedit and radar graphs

Keywords

Neutralizing antibodies; HIV epitopes; Gp41 mutations; Vaccine targets; Bioedit and radar graphs

Introduction

Human Immunodeficiency virus -1 (HIV) causes Acquired Immuno Deficiency Syndrome (AIDS) and the prevalence of the syndrome always increases with almost 6500 new infections every day [1]. The failure to design a successful prophylactic vaccine for about three decades, ever since the discovery of the etiological agent is burdening the field of medicine [2]. The enhancement of the knowledge of the immune response and the viral pathogenesis with its complex genome organization remains as a need across the decades. The quality of any successful vaccine is the elicitation of the neutralizing antibody against a specific pathogen. In the event of the viral envelope glycoprotein being conserved, those epitopes can be used successfully in conventional vaccine strategies as with the Hepatitis B Virus vaccine licensed for human use [3]. Subsequently, such strategies have been improvised by the recombinant-DNA technology and efforts for an edible vaccine for HBV [4]. Such conserved epitopes which can elicit neutralizing antibody response to prevent the establishment of HIV infection are needed to address the prevailing epidemiology. The current vaccine design efforts are aimed at producing broadly neutralizing antibodies that can be improved for achieving the long awaited goal [5]. The 2F5 and 4E10 epitopes have been evaluated for their potency to elicit neutralizing antibodies in combination with the other epitopes of HIV [6].
Passive immunization with neutralizing antibodies has demonstrated pre-exposure immunity by the host by preventing the establishment of the infection in primate models [7]. The mucosal challenge with the virus after passive immunization using combinations of human monoclonal antibodies were conferring protection in primate models [8]. The regions that were targeted about a decade ago were more capable of cross-clade neutralization and were conferring hope for a prophylactic or a therapeutic vaccine [9]. The hydrophobicity that influences the reactivity of the 2F5 antibody with its complementarity determining regions has been studied in order to enhance the ability of these antibodies to neutralize [10].
These 2F5 and 4E10 monoclonal antibodies targeted to the epitopes in the MPER of the gp41 subunit interfere with the fusion of the viral envelope and the host cell membrane. Refined atomic level interactions of 2F5 epitope-paratope were studied for preventing the viral entry into the host cells [11]. The monoclonal antibody targeting the purported conserved epitopes was used alone and in combination to study their ability to neutralize the virus within and across the clades by testing on primate models by intravenous and mucosal immunization [12]. The non- neutralizing antibodies that conjoin with the 2F5 monoclonal antibody in blocking the HIV-1 entry were also reported with their crystal structures [13]. With these prior data and knowledge, the 2F5 and 4E10 antibodies targeted epitopes, so far have been the putative vaccine targets. On the other hand, the HIV has evolved multiple strategies to avoid the neutralization by these antibodies that target these regions of interest in the envelope glycoprotein [14].
The vaccine that is not achieved across the decades is due to the challenges featured by the genome organization of the retrovirus. The dynamic replication capacity and the erroneous patterns of the recombining reverse transcription attribute to the evasion of the virus from surveillance by these neutralizing antibodies. These are the important implications to be considered in vaccine strategies [15]. In this paper, we present the mutations in the targets of 2F5 and 4E10 antibodies in the sequences from the different parts of the world to understand the evolutionary trend that favors the loss of conservation to evade the neutralization by these putative vaccine candidates.

Methods

Sequences reported in the Pubmed spanning almost all regions of the world were retrieved randomly and stored in FASTA format (n=1517). The sequences were then aligned using the ClustalW feature of the Bioedit V7.0.9.0 and the consensus across the 1517 sequences was generated. The conservation plot mode was used to view the substitution and the changes across the sequences. The DNA sequences were then toggle translated to obtain the corresponding protein sequences. The positions corresponding to the two epitopes of the 2F5 and 4E10 neutralizing antibodies were extracted and exported as an XML file. The analyses were done using Microsoft Excel 2007. The radar graphs were constructed using the charting features of MSExcel 2007 to depict the substitutions with their extent and patterns.

Results

The proportions of HIV gp41 sequences that were reported from different countries across the world are represented in the Figure 1. Each residue in both of the targets is found to be substituted by one or more amino acid. The substitutions and their patterns found in each residue of the two epitopes of 2F5 and 4E10 antibodies are represented here in Figures 2 and 3 respectively.
Figure 1: The metadata (from the Pubmed) taken for this investigation was reported from different countries and the proportion of the sequences from each country is represented pictorially.

Discussion

The substitutions found in the targets of the 2F5 and 4E10 antibodies can critically impact on the antibody affinity or specificity. The glutamine E38 has been notably substituted by Alanine. This substitution of an acidic amino acid by alanine (an aliphatic amino acid) can vitally influence the antibody affinity. The prevalence of substitution at K41 is found in some sequences and it is evident from the distortion of the circle in the Figure 2d whereas the degree of conservation is higher in the D39 and W42 residues and it is evident in the perfection of the circle in Figures 2c and 2e. The failure of complex formation due to the replacement of K41 by glutamine and alanine has been demonstrated. The substitutions in the neighboring residues also had resulted in the lack of binding. This change in themolecular environment due to the alteration at the DKW residues of the core epitope confers resistance to the respective antibody as the DKW motif centers the β-turn conformation of the gp41 in the MPER [16]. Also the A43 has been changed or deleted in a few sequences while the S44 has been substituted by N44 which can influence on the 2F5 antibody interaction due to the change in the side chain as a result of the substitution.
Figure 2: 2a) E38 is substituted as A38 to a notable extent. 2b) L39 is not significantly sybstituted. 2c) D40 is not significantly substituted. 2d) K41 is having a minor proportion of substitution. 2e) W42 is not having a significant substitutions. 2f) A43 has a notable proportion of substitutions. 2g) S44 has been notably substituted as N44.
Pertaining to the 4E10 antibody target, the occurrence of mutations as depicted in Figure 3 may potentially affect its neutralizing ability. Sequences have substitutions at all the residues except W48 and F49 that are contained in the tryptophan rich region. Crystal structure of the 4E10 antibody has been reported in conjugation with its epitope (NWFDIT) [17]. Alanine substitution within or near the epitope has shown to decrease the affinity of the 2F5 antibody to its epitope [18]. The residue D50 is tended to change to N50 and S50. This is evident in the shift of the base of the radiating arm moving towards N from D (Figure 3d). This kind of substitution or its absence enables the virus to modulate the glycosylation ability. The glycan shield can mask the virus from being recognized by the 4E10 antibodies [19]. Such glycosylation feature greatly enhances the viral entry and replication in the macrophages [20]. The T52 has been substituted as S52 and the substitution will not greatly change the reactivity as both these amino acids have the same side chain. These mutations resulting in drastic side chain changes of the amino acids may render the target inaccessible or inert for the antibodies to react. These neutralizing antibodies vary in their ability of interaction with the targeted conserved regions and with the functional conformations of the glycoprotein trimers to prevent the entry of the HIV-1 into the host cells [21].
Figure 3: 3a) N47 is substituted as S47 to a notable extent. 3b) W48 is very negligibly substituted 3c) F49 is having a negligible proportion of substitution. Stop codons were also found as substitutes in a few sequences 3d) N50 is substituted mostly as D50 and S50. Frame shift or deletions were also observed in a few sequences. 3e) I51 has few substitutions. 3f) T52 is substituted as S52 and has been deleted in a few sequences.
Mutations occurring on this putative vaccine target (as in the figures 2 and 3) are plausibly due to the pressure of the immune system that targets the epitope with the circulating neutralizing antibodies or they might be drug induced. The diverse conformations of the glycoproteins arising due to the mutations can result in the masking of the virus to escape neutralization [22]. It has been demonstrated that no single substitution will confer resistance to the 2F5 or 4E10 antibodies. The synergy of the 2F5 and 4E10 antibodies was not observed, but the 2F5 was antagonized by 4E10 [18]. The stoichiometric estimation of the neutralizing ability based on the number of trimers that are found on a virion as a key influencing factor has also been investigated for improving the knowledge of the virus entry [23].
RNA genome of viruses as an entity in the rapidly evolving RNA biospheres have higher rates of mutations and are evolutionarily competent against their hosts [24]. HIV as a member of the Retroviridae family stands as a classic example of this remarkable evolutionary competence [25]. This enormous genetic diversity of the HIV genome with its adaptability complemented by the error prone or the ‘infidel’ reverse transcriptase helps the virus to replicate at a higher rate [26]. In an in vivo milieu, the mutations that are advantageous for the virus can be selected and copied according to the copy choice model that explains recombination in HIV favoring survivable variants [27]. This strategy of recombination can exponentially add to the evolutionary competency of the retrovirus [28]. Hence, with the observed patterns of substitutions in the 2F5 and 4E10 targets, we suggest the need for a better immunogenic epitope that is essentially conserved for a prophylactic vaccine candidature or a therapeutic agent that can promise a way forward in combating HIV infection.

Acknowledgments

The authors thank the University Grants Commission of India for funding the project.

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