Structural Descriptors of gp120 V3 Loop for the Prediction of HIV-1 Coreceptor Usage

HIV-1 cell entry commonly uses, in addition to CD4, one of the chemokine receptors CCR5 or CXCR4 as coreceptor. Knowledge of coreceptor usage is critical for monitoring disease progression as well as for supporting therapy with the novel drug class of coreceptor antagonists. Predictive methods for inferring coreceptor usage based on the third hypervariable (V3) loop region of the viral gene coding for the envelope protein gp120 can provide us with these monitoring facilities while avoiding expensive phenotypic tests. All simple heuristics (such as the 11/25 rule) as well as statistical learning methods proposed to date predict coreceptor usage based on sequence features of the V3 loop exclusively. Here, we show, based on a recently resolved structure of gp120 with an untruncated V3 loop, that using structural information on the V3 loop in combination with sequence features of V3 variants improves prediction of coreceptor usage. In particular, we propose a distance-based descriptor of the spatial arrangement of physicochemical properties that increases discriminative performance. For a fixed specificity of 0.95, a sensitivity of 0.77 was achieved, improving further to 0.80 when combined with a sequence-based representation using amino acid indicators. This compares favorably with the sensitivities of 0.62 for the traditional 11/25 rule and 0.73 for a prediction based on sequence information as input to a support vector machine and constitutes a statistically significant improvement. A detailed analysis and interpretation of structural features important for classification shows the relevance of several specific hydrogen-bond donor sites and aliphatic side chains to coreceptor specificity towards CCR5 or CXCR4. Furthermore, an analysis of side chain orientation of the specificity-determining residues suggests a major role of one side of the V3 loop in the selection of the coreceptor. The proposed method constitutes the first approach to an improved prediction of coreceptor usage based on an original integration of structural bioinformatics methods with statistical learning

Clustering of HIV-1 Subtypes Based on gp120 V3 Loop electrostatic properties

<p>Abstract</p> <p>Background</p> <p>The V3 loop of the glycoprotein gp120 of HIV-1 plays an important role in viral entry into cells by utilizing as coreceptor CCR5 or CXCR4, and is implicated in the phenotypic tropisms of HIV viruses. It has been hypothesized that the interaction between the V3 loop and CCR5 or CXCR4 is mediated by electrostatics. We have performed hierarchical clustering analysis of the spatial distributions of electrostatic potentials and charges of V3 loop structures containing consensus sequences of HIV-1 subtypes.</p> <p>Results</p> <p>Although the majority of consensus sequences have a net charge of +3, the spatial distribution of their electrostatic potentials and charges may be a discriminating factor for binding and infectivity. This is demonstrated by the formation of several small subclusters, within major clusters, which indicates common origin but distinct spatial details of electrostatic properties. Some of this information may be present, in a coarse manner, in clustering of sequences, but the spatial details are largely lost. We show the effect of ionic strength on clustering of electrostatic potentials, information that is not present in clustering of charges or sequences. We also make correlations between clustering of electrostatic potentials and net charge, coreceptor selectivity, global prevalence, and geographic distribution. Finally, we interpret coreceptor selectivity based on the N⁶X⁷T⁸|S⁸X⁹sequence glycosylation motif, the specific positive charge location according to the 11/24/25 rule, and the overall charge and electrostatic potential distribution.</p> <p>Conclusions</p> <p>We propose that in addition to the sequence and the net charge of the V3 loop of each subtype, the spatial distributions of electrostatic potentials and charges may also be important factors for receptor recognition and binding and subsequent viral entry into cells. This implies that the overall electrostatic potential is responsible for long-range recognition of the V3 loop with coreceptors CCR5/CXCR4, whereas the charge distribution contributes to the specific short-range interactions responsible for the formation of the bound complex. We also propose a scheme for coreceptor selectivity based on the sequence glycosylation motif, the 11/24/25 rule, and net charge.</p

Computational approaches for improving treatment and prevention of viral infections

The treatment of infections with HIV or HCV is challenging. Thus, novel drugs and new computational approaches that support the selection of therapies are required. This work presents methods that support therapy selection as well as methods that advance novel antiviral treatments. geno2pheno[ngs-freq] identifies drug resistance from HIV-1 or HCV samples that were subjected to next-generation sequencing by interpreting their sequences either via support vector machines or a rules-based approach. geno2pheno[coreceptor-hiv2] determines the coreceptor that is used for viral cell entry by analyzing a segment of the HIV-2 surface protein with a support vector machine. openPrimeR is capable of finding optimal combinations of primers for multiplex polymerase chain reaction by solving a set cover problem and accessing a new logistic regression model for determining amplification events arising from polymerase chain reaction. geno2pheno[ngs-freq] and geno2pheno[coreceptor-hiv2] enable the personalization of antiviral treatments and support clinical decision making. The application of openPrimeR on human immunoglobulin sequences has resulted in novel primer sets that improve the isolation of broadly neutralizing antibodies against HIV-1. The methods that were developed in this work thus constitute important contributions towards improving the prevention and treatment of viral infectious diseases.Die Behandlung von HIV- oder HCV-Infektionen ist herausfordernd. Daher werden neue Wirkstoffe, sowie neue computerbasierte Verfahren benötigt, welche die Therapie verbessern. In dieser Arbeit wurden Methoden zur Unterstützung der Therapieauswahl entwickelt, aber auch solche, welche neuartige Therapien vorantreiben. geno2pheno[ngs-freq] bestimmt, ob Resistenzen gegen Medikamente vorliegen, indem es Hochdurchsatzsequenzierungsdaten von HIV-1 oder HCV Proben mittels Support Vector Machines oder einem regelbasierten Ansatz interpretiert. geno2pheno[coreceptor-hiv2] bestimmt den HIV-2 Korezeptorgebrauch dadurch, dass es einen Abschnitt des viralen Oberflächenproteins mit einer Support Vector Machine analysiert. openPrimeR kann optimale Kombinationen von Primern für die Multiplex-Polymerasekettenreaktion finden, indem es ein Mengenüberdeckungsproblem löst und auf ein neues logistisches Regressionsmodell für die Vorhersage von Amplifizierungsereignissen zurückgreift. geno2pheno[ngs-freq] und geno2pheno[coreceptor-hiv2] ermöglichen die Personalisierung antiviraler Therapien und unterstützen die klinische Entscheidungsfindung. Durch den Einsatz von openPrimeR auf humanen Immunoglobulinsequenzen konnten Primersätze generiert werden, welche die Isolierung von breit neutralisierenden Antikörpern gegen HIV-1 verbessern. Die in dieser Arbeit entwickelten Methoden leisten somit einen wichtigen Beitrag zur Verbesserung der Prävention und Therapie viraler Infektionskrankheiten

Analysis of HIV-host interaction on different scales

The human immunodeficiency virus depends on molecular pathways of the host for efficient replication and spread. The intricate network of host-virus interactions shapes the virus\u27; evolution by driving the pathogen to evade immune recognition and constraining it to maintain its capacity to replicate. Study of the HIV-host interactions provides important insights into viral evolution, pathogenicity and potential treatment strategies. This thesis presents an analysis of HIV-host interactions on several scales, ranging from individual protein interactions to whole genomes. On the scale of individual interaction we analyze structural and physical determinants of the interaction between host TRIM5alpha and virus capsid — an interaction of potential therapeutic interest due to the capacity of TRIM5alpha to block retroviral infections. On the scale of viral population we present two studies of a highly variable region of the virus genome involved in the interaction with host cell coreceptors upon virus cell entry. The studies provide insights into the virus evolution and the physicochemical and structural properties related to its interaction with cellular coreceptors. On the scale of the single cell we develop models of HIV cell entry involving virus, host and environmental factors. The models represent a comprehensive picture of the virus phenotype and allow one to view the variability of virus phenotypes on 2D phenotype maps. On the genomic scale we perform a large-scale analysis of all HIV-host interactions. This study reveals insights into general patterns of the host-pathogen evolution and suggests candidate host proteins involved in interactions potentially important for the infection and interesting for further study on other scales. Interactions and processes crucial for the HIV infection reemerge across the scales pointing to the importance of integrative, multi-scale studies of host-pathogen biology.Das Humane Immundefizienz-Virus hängt von molekularen Mechanismen des Wirts für seine effiziente Replikation und Ausbreitung ab. Das komplizierte Netzwerk von Wirt-Virus Interaktionen formt die Evolution des Virus, indem es den Erreger dazu bringt, sich der Erkennung durch das Immunsystem zu entziehen und seine Replikationskapazität aufrecht zu erhalten. Das Studium der HIV-Wirt Interaktionen erlaubt wichtige Einblicke in die viralen Evolution, die Pathogenität des Virus, sowie mögliche Behandlungsstrategien. Diese Arbeit stellt eine Analyse der HIV-Wirt-Interaktionen in mehreren Größenordnungen vor, von einzelnen Protein-Interaktionen bis hin zur Analyse ganzer Genome. In Hinblick auf einzelne Interaktionen untersuchen wir strukturelle und physikalische Determinanten der Interaktion zwischen dem Wirtfaktor TRIM5alpha; und dem viralen Kapsid - eine Interaktion, die von therapeutischem Interesse ist wegen der Fähigkeit von TRIM5alpha, retrovirale Infektionen zu blockieren. In Hinblick auf virale Populationen präsentieren wir zwei Studien einer hochvariablen Region des viralen Genoms, die in der Interaktion des Virus mit zellulären Rezeptoren des Wirts beim viralen Zelleintritt involviert sind. Diese Studien geben Einblick in die virale Evolution und die physikalisch-chemischen und strukturellen Eigenschaften des Virus bezüglich dessen Interaktion mit zellulären Ko-Rezeptoren. Auf der Skala der einzelnen Zelle entwickeln wir Modelle des HIV Zelleintritts welche das Virus, den Wirt und Umgebungsfaktoren berücksichtigen. Diese Modelle bieten ein umfassendes Bild des viralen Phänotyps und erlauben es, die Variabilität des Virus auf 2D-Phänotyp-Karten zu visualisieren. Im genomweiten Maßstab führen wir eine groß angelegte Analyse aller HIV-Wirt-Interaktionen durch. Diese Studie erlaubt Einblicke in allgemeine Muster der Wirt-Pathogen-Evolution und identifiziert Kandidaten für Wirtsproteine, deren Interaktionen potenziell wichtig für die virale Infektion sind und deren weitere Untersuchung in anderen Größenordnungen von Interesse ist. Interaktionen und Prozesse, die von entscheidender Bedeutung für die HIV-Infektion sind zeigen sich wiederholt in allen untersuchten Maßstäben und unterstreichen die Bedeutung einer integrativen und multi-skalaren Untersuchung der Wirt-Pathogen-Biologie

In silico Analysis of HIV-1 Env-gp120 Reveals Structural Bases for Viral Adaptation in Growth-Restrictive Cells

Variable V1/V2 and V3 loops on human immunodeficiency virus type 1 (HIV-1) envelope-gp120 core play key roles in modulating viral competence to recognize two infection receptors, CD4 and chemokine-receptors. However, molecular bases for the modulation largely remain unclear. To address these issues, we constructed structural models for a full-length gp120 in CD4-free and -bound states. The models showed topologies of gp120 surface loop that agree with those in reported structural data. Molecular dynamics simulation showed that in the unliganded state, V1/V2 loop settled into a thermodynamically stable arrangement near V3 loop for conformational masking of V3 tip, a potent neutralization epitope. In the CD4-bound state, however, V1/V2 loop was rearranged near the bound CD4 to support CD4 binding. In parallel, cell-based adaptation in the absence of anti-viral antibody pressures led to the identification of amino acid substitutions that individually enhance viral entry and growth efficiencies in association with reduced sensitivity to CCR5 antagonist TAK-779. Notably, all these substitutions were positioned on the receptors binding surfaces in V1/V2 or V3 loop. In silico structural studies predicted some physical changes of gp120 by substitutions with alterations in viral replication phenotypes. These data suggest that V1/V2 loop is critical for creating a gp120 structure that masks co-receptor binding site compatible with maintenance of viral infectivity, and for tuning a functional balance of gp120 between immune escape ability and infectivity to optimize HIV-1 replication fitness

Structural Modeling of HIV-1 Env-gp120

Variable V1/V2 and V3 loops on human immunodeficiency virus type 1 (HIV-1) envelope-gp120 core play key roles in modulating viral competence to recognize two infection receptors, CD4 and chemokine-receptors. However, molecular bases for the modulation largely remain unclear. To address these issues, we constructed structural models for a full-length gp120 in CD4-free and -bound states. The models showed topologies of gp120 surface loop that agree with those in reported structural data. Molecular dynamics simulation showed that in the unliganded state, V1/V2 loop settled into a thermodynamically stable arrangement near V3 loop for conformational masking of V3 tip, a potent neutralization epitope. In the CD4-bound state, however, V1/V2 loop was rearranged near the bound CD4 to support CD4 binding. In parallel, cell-based adaptation in the absence of anti-viral antibody pressures led to the identification of amino acid substitutions that individually enhance viral entry and growth efficiencies in association with reduced sensitivity to CCR5 antagonist TAK-779. Notably, all these substitutions were positioned on the receptors binding surfaces in V1/V2 or V3 loop. In silico structural studies predicted some physical changes of gp120 by substitutions with alterations in viral replication phenotypes. These data suggest that V1/V2 loop is critical for creating a gp120 structure that masks co-receptor binding site compatible with maintenance of viral infectivity, and for tuning a functional balance of gp120 between immune escape ability and infectivity to optimize HIV-1 replication fitness

Experimental Determination of the Topology of the HIV-1 gp41 C-Terminal Tail

The C-terminal tail (CTT) of HIV gp41 has been traditionally viewed as a cytoplasmic domain. Genetic studies demonstrating functional interactions between the CTT and various intracellular partners have implicitly reinforced this view. However, antibody neutralization data and biochemical studies have suggested that the CTT is, or can be, externally localized under certain condition. Additionally, other studies have demonstrated that the CTT is dispensable for in vitro virus replication. After nearly three decades of HIV research, the function and structure of the CTT remain elusive. Our goals, then, were twofold: (i) to determine the overall conservation of the CTT in an attempt to provide an understanding of the functional and structural relevance of the CTT; and, (ii) to provide an experimental topological map of the CTT in an attempt to understand and align observed CTT topology(ies) with the functional necessity of a cytoplasmic CTT. We believe that we made significant contributions to the understanding of CTT topology and its relationship to current published functional studies. The initial studies demonstrated that the CTT sequence is conserved at a level that is intermediate between the highly variable gp120 region and the relatively conserved gp41 ectodomain. Additionally, physicochemical and structural properties of CTT sequences were found to be conserved in spite of the relatively high sequencevariability. These studies demonstrated for the first time that the CTT sequence, while highly variable, contains highly conserved structural and chemical properties that suggest a functional requirement for the CTT. Topology studies of the CTT indicated that the topology of the CTT can be distinct between the surface of Env-expressing cells and viral particles. Additionally, dynamic rearrangement of the CTT was observed as a function of antibody neutralization. These findings prompted a theoretical study of gp41 CTT predicted topology and the proposal of a topological model that we believe is consistent with all published studies regarding the localization of the CTT

Highly Accurate Structure-Based Prediction of HIV-1 Coreceptor Usage Suggests Intermolecular Interactions Driving Tropism

HIV-1 entry into host cells is mediated by interactions between the V3-loop of viral glycoprotein gp120 and chemokine receptor CCR5 or CXCR4, collectively known as HIV-1 coreceptors. Accurate genotypic prediction of coreceptor usage is of significant clinical interest and determination of the factors driving tropism has been the focus of extensive study. We have developed a method based on nonlinear support vector machines to elucidate the interacting residue pairs driving coreceptor usage and provide highly accurate coreceptor usage predictions. Our models utilize centroid-centroid interaction energies from computationally derived structures of the V3-loop:coreceptor complexes as primary features, while additional features based on established rules regarding V3-loop sequences are also investigated. We tested our method on 2455 V3-loop sequences of various lengths and subtypes, and produce a median area under the receiver operator curve of 0.977 based on 500 runs of 10-fold cross validation. Our study is the first to elucidate a small set of specific interacting residue pairs between the V3-loop and coreceptors capable of predicting coreceptor usage with high accuracy across major HIV-1 subtypes. The developed method has been implemented as a web tool named CRUSH, CoReceptor USage prediction for HIV-1, which is available at http://ares.tamu.edu/CRUSH/

POPISK: T-cell reactivity prediction using support vector machines and string kernels

BACKGROUND: Accurate prediction of peptide immunogenicity and characterization of relation between peptide sequences and peptide immunogenicity will be greatly helpful for vaccine designs and understanding of the immune system. In contrast to the prediction of antigen processing and presentation pathway, the prediction of subsequent T-cell reactivity is a much harder topic. Previous studies of identifying T-cell receptor (TCR) recognition positions were based on small-scale analyses using only a few peptides and concluded different recognition positions such as positions 4, 6 and 8 of peptides with length 9. Large-scale analyses are necessary to better characterize the effect of peptide sequence variations on T-cell reactivity and design predictors of a peptide's T-cell reactivity (and thus immunogenicity). The identification and characterization of important positions influencing T-cell reactivity will provide insights into the underlying mechanism of immunogenicity. RESULTS: This work establishes a large dataset by collecting immunogenicity data from three major immunology databases. In order to consider the effect of MHC restriction, peptides are classified by their associated MHC alleles. Subsequently, a computational method (named POPISK) using support vector machine with a weighted degree string kernel is proposed to predict T-cell reactivity and identify important recognition positions. POPISK yields a mean 10-fold cross-validation accuracy of 68% in predicting T-cell reactivity of HLA-A2-binding peptides. POPISK is capable of predicting immunogenicity with scores that can also correctly predict the change in T-cell reactivity related to point mutations in epitopes reported in previous studies using crystal structures. Thorough analyses of the prediction results identify the important positions 4, 6, 8 and 9, and yield insights into the molecular basis for TCR recognition. Finally, we relate this finding to physicochemical properties and structural features of the MHC-peptide-TCR interaction. CONCLUSIONS: A computational method POPISK is proposed to predict immunogenicity with scores which are useful for predicting immunogenicity changes made by single-residue modifications. The web server of POPISK is freely available at http://iclab.life.nctu.edu.tw/POPISK

Examination of the role of envelope directed antibodies on co-receptor usage in HIV-1B infection

HIV-1 primarily utilizes the CCR5 receptor as a co-receptor, but over time, viruses can evolve to use the CXCR4 protein. Changes in the viral envelope V3 loop mediate this switch. The emergence of CXCR4-utilizing viruses has been presumed to occur as a consequence of decreased humoral immunity. We show that exclusively CXCR4-using (X4) viruses contain a 2 to 3 amino acid insertion in the V3 loop. Structural modeling revealed that this insertion caused a protrusion in the V3 loop, which impacts CCR5 receptor interaction. These genotypic and structural motifs affected neutralization susceptibility because X4, as compared to co-circulating CCR5-utilizing (R5) viruses, were less neutralization sensitive to autologous contemporaneous and heterologous plasma. Individuals with co-circulating X4 and R5, as compared to those with only R5, viruses had similar neutralization breadth and potency indicating that the emergence of X4 viruses is not associated with decreased humoral immunity. These results suggest that X4 viruses are neutralization escape variants and arise due to humoral selective pressure. This work has implications for future antibody-based therapeutics. Along with providing a framework for developing an HIV-1 vaccine, broadly neutralizing antibodies (bnAbs) are also being investigated as a potential therapeutic. BnAbs target a limited number of conserved HIV-1 envelope structures, including glycans in and around the V1/V2 and V3 domains. Along with the V3 loop, changes in V1/V2 are also known to impact co-receptor usage. We show that viruses that exclusively use the CXCR4 co-receptor, as compared to variants that only utilize CCR5, were less neutralization sensitive to V1/V2 and V3 directed bnAbs. In contrast, R5 and X4 viruses did not demonstrate neutralization differences to bnAbs that target non-V1/V2 and V3 envelope regions, such as the CD4 binding site and the membrane proximal external region. Structural modeling revealed that the predicted orientation of the V1/V2 loop among diverse HIV-1 variants predicts susceptibility to V3 loop directed bnAbs. In aggregate, our results suggest that viruses with different co-receptor usage have differing bnAb susceptibility. Furthermore, structural modeling may be used as a tool to predict neutralization susceptibility to bnAbs against regions associated with co-receptor usage.2020-06-12T00:00:00