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

Studies of Protein Interactions and Knowledge-Based Drug Design: (A) The Electrostatic Nature of Recognition Between HIV-1 gp120 V3 Loop and Coreceptors CCR5/CXCR4, (B) Complement System Inhibition by Compstatin Family Peptides

Computational and experimental methods were used to understand (i) protein interactions involving the V3 loop of gp120 of HIV-1 with coreceptors in host cells and (ii) peptide analogs from the compstatin family to human C3. Computational methods, including molecular dynamics (MD) simulations and electrostatic calculations, provide quantitative predictions of dynamics and interactions at atomic resolution, while experimental methods, including surface plasmon resonance (SPR) and enzyme-linked immunosorbent assays (ELISA) are needed to confirm binding and inhibition. HIV-1 entry into host cells is mediated by the interaction of the V3 loop of gp120 and coreceptors CCR5 or CXCR4 on host cell surfaces, with assistance of viral protein gp41 and cell receptor CD4. The mechanism of coreceptor selectivity is not well understood, given the sequence variability and structural flexibility of the V3 loop. Positive net charge is a persistent physicochemical property throughout HIV subtypes and has been recognized as an influencing factor for cell entry. Electrostatic analyses of V3 loop structures with consensus sequences from HIV subtypes, show similar electrostatic potential characteristics, irrespective of sequence variability. Charge and other sequence-based criteria were combined to develop a scheme for determining coreceptor selection. In addition, MD simulations provide insight into loop dynamics, indicating that persistent salt bridges contribute in keeping the two loop strands in proximity, therefore providing a charged scaffold for electrostatic interactions with coreceptors, irrespective of structural variability. Compstatin family peptides are inhibitors of the complement system and potential drug candidates against autoimmune and inflammatory diseases. Compstatin analogs are cyclic peptides that inhibit cleavage of human C3, therefore preventing further complement system activation. Introduction of tryptophan residues at the termini resulted in potent analogs, but suffering from reduced solubility. To balance hydrophobicity (important for binding) and polarity (important for solubility), additional analogs were designed guided by MD simulation results of bound analogs to C3. New analogs with polar substitutions at the N-terminus, including dipeptide sequence extensions and use of methylated tryptophan residues, were experimentally tested with ELISAs, demonstrating comparable inhibition to that of known analogs, but with improved solubility

Insights into the Structure, Correlated Motions, and Electrostatic Properties of Two HIV-1 gp120 V3 Loops

The V3 loop of the glycoprotein 120 (gp120) is a contact point for cell entry of HIV-1 leading to infection. Despite sequence variability and lack of specific structure, the highly flexible V3 loop possesses a well-defined role in recognizing and selecting cell-bound coreceptors CCR5 and CXCR4 through a mechanism of charge complementarity. We have performed two independent molecular dynamics (MD) simulations to gain insights into the dynamic character of two V3 loops with slightly different sequences, but significantly different starting crystallographic structures. We have identified highly populated trajectory-specific salt bridges between oppositely charged stem residues Arg9 and Glu25 or Asp29. The two trajectories share nearly identical correlated motions within the simulations, despite their different overall structures. High occupancy salt bridges play a key role in the major cross-correlated motions in both trajectories, and may be responsible for transient structural stability in preparation for coreceptor binding. In addition, the two V3 loops visit conformations with similarities in spatial distributions of electrostatic potentials, despite their inherent flexibility, which may play a role in coreceptor recognition. It is plausible that cooperativity between overall electrostatic potential, charged residue interactions, and correlated motions could be associated with a coreceptor selection and binding

Kinetics And Thermodynamics Of Dbpa Protein\u27S C-Terminal Domain Interaction With Rna

DbpA is an Escherichia coli DEAD-box RNA helicase implicated in RNA structural isomerization in the peptide bond formation site. In addition to the RecA-like catalytic core conserved in all of the members of DEAD-box family, DbpA contains a structured C-terminal domain, which is responsible for anchoring DbpA to hairpin 92 of 23S ribosomal RNA during the ribosome assembly process. Here, surface plasmon resonance was used to determine the equilibrium dissociation constant and the microscopic rate constants of the DbpA C-terminal domain association and dissociation to a fragment of 23S ribosomal RNA containing hairpin 92. Our results show that the DbpA protein\u27s residence time on the RNA is 10 times longer than the time DbpA requires to hydrolyze one ATP. Thus, our data suggest that once bound to the intermediate ribosomal particles via its RNA-binding domain, DbpA could unwind a number of double-helix substrates before its dissociation from the ribosomal particles

Side Chain Interaction Energies (in kcal/mol) for 2B4C (A) and 2QAD (B).

<p>Energies were computed and averaged over the trajectory, as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049925#s2" target="_blank">Methods</a>. The blue, red, and green bars correspond to non-polar, polar and total interaction energies, respectively. The most representative structures throughout the trajectories (global minimum; black circle in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049925#pone-0049925-g007" target="_blank">Figure 7</a>, below) are shown in panel (C) for 2B4C and in panel (D) for 2QAD. The backbone is shown in tube representation and side chains are shown in stick representation. Negatively and positively charged residues involved in salt bridges are shown in red and blue, respectively, and disulfide bridge residues in yellow. Salt bridges and β-bridges are marked with dashed lines. The backbone of the base (residues 1–4, 31–35), stem (residues 5–10, 21–30) and tip (residues 11–20) regions is colored in cyan, black and purple color, respectively. The rest of the side chains are shown in thin pink licorice representation. Hydrogen atoms are omitted for clarity.</p

Molecular model of the V3 loops for 2B4C (A) and 2QAD (B).

<p>Backbone is shown in tube representation. The base (residues 1–4, 31–35), stem (residues 5–10, 21–30) and tip (residues 11–20) regions are colored in cyan, black and purple color, respectively. Specific side chain residues are shown and are color-coded: yellow denotes residues 1 and 35, forming the disulfide bridge; blue denotes residues involved in the glycosylation motif; red denotes residues involved in the “11/24/25” rule; green denotes the conserved GPG motif at the tip of the loop; orange denotes residues involved in salt bridges within the present MD simulations, including residue 9 (blue in 2B4C and 2QAD) and residue 25 (red in 2QAD).</p

Occupancy of salt bridges throughout the trajectories. Salt bridges were calculated with a cutoff of 5 Å.

<p>Occupancy of salt bridges throughout the trajectories. Salt bridges were calculated with a cutoff of 5 Å.</p

Charged Interactions within Principal Component 1 for 2B4C (A) and 2QAD (B).

<p>Axes denote the residue number in sequence. Colors correspond to those of the “extreme” structures observed during the principal component 1 of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049925#pone-0049925-g005" target="_blank">Figure 5</a>.</p