The Conformation and Orientation of a 27-Residue CCR5 Peptide in a Ternary Complex with HIV-1 gp120 and a CD4-Mimic Peptide

Abstract

Interaction of CCR5 with the HIV-1 gp120-CD4 complex involves its amino-terminal domain (Nt-CCR5) and requires sulfation of 2-4 tyrosine residues in Nt-CCR5. The conformation of a 27-residue Nt-CCR5 peptide, sulfated at Y10 and Y14, was studied in both its free form and in a ternary complex with deglycosylated-gp120 and a CD4-mimic peptide. NMR experiments revealed a helical conformation at the center of Nt-CCR5(1-27) which is induced upon gp120 binding, as well as a helical propensity for the free peptide. A well-defined structure for the bound peptide was determined for residues 7-23, increasing by two-fold the length of Nt-CCR5's known structure. Two-dimensional saturation transfer experiments and measurement of relaxation-times highlighted Nt-CCR5 residues Y3, V5, P8-T16, E18, I23 and possibly D2 as the main binding determinant. A calculated docking model for Nt-CCR5(1-27) suggests that residues 2-22 of Nt-CCR5 interact with the bases of V3 and C4 while the C-terminal segment of Nt-CCR5(1-27) points towards the target cell membrane reflecting an Nt-CCR5 orientation that differs by 180° from a previous model. A gp120 site that could accommodate CCR5Y3 in a sulfated form has been identified. The present model attributes a structural basis for binding interactions to all gp120 residues previously implicated in Nt-CCR5 binding. Moreover, the strong interaction of sulfated CCR5Tyr14 with gp120Arg440 revealed by the model and the previously found correlation between E322 and R440 mutations shed light on the role of these residues in HIV-1 phenotype conversion furthering our understanding of CCR5 recognition by HIV-1.

Peptides from Second Extracellular Loop of C-C Chemokine Receptor Type 5 (CCR5) Inhibit Diverse Strains of HIV-1

“…At this pH, we did not observe oxidation to the dimeric form of the Cys-containing peptides as detected by HR-MS. The CCR5 N-terminal peptide comprising residues 2-18 was synthesized using previously reported procedures (20).…”

“…In the case of the CCR5 N terminus, this tactic has proven especially useful for defining post-translational modifications and peptide sequences critical for function as well as inhibition of HIV-1 entry (15)(16)(17)(18). They also facilitated determination of high-resolution structures of CCR5 N terminus peptides in complex with gp120, furthering our understanding of this interaction at an atomic level (19,20) and making possible the discovery of small molecule CCR5 N terminus mimetics (21). Such studies demonstrate the value in using peptide fragments of G protein-coupled receptors as tools to study coreceptor interactions because appropriate peptides tend to function in part as coreceptor mimics (22,23).…”

Incompatible Natures of the HIV-1 Envelope in Resistance to the CCR5 Antagonist Cenicriviroc and to Neutralizing Antibodies

“…CCR5 antagonists are suggested to block HIV-1 infection by means of a structural change in CCR5, which prevents interaction between HIV-1 and the CCR5 antagonist-bound form of CCR5 (2)(3)(4)(5). The V3 loop and CD4i region of HIV-1 Env are important for interaction with CCR5 and therefore play a key role in resistance to CCR5 antagonists.…”

“…Most CCR5 antagonists reported so far bind a hydrophobic pocket of CCR5, causing a structural rearrangement of extracellular loop 2 and the N-terminal region, which interact with HIV-1 Env glycoprotein (2)(3)(4)(5). Many CCR5 antagonists, such as TAK-779 (6), TAK-220 (7), vicriviroc (VCV) (8), aplaviroc (9), and maraviroc (MVC) (10), have been developed, yet only MVC is clinically used for the treatment of HIV-1-infected patients.…”

Short Communication: Synergism Between a CD4-Mimic Peptide and Antibodies Elicited by a Constrained V3 Peptide

“…CD4M33, i.e., TpaNLHFCQLRCKSLGLLGK CAGSBipCACV-NH 2 (Tpa, thiopropionic acid; Bip, biphenylalanine), was synthesized as described previously. 18,25 For each strain, neutralization was evaluated for the serum alone starting at a 1:10 serum dilution followed by consecutive 2-fold dilutions. Similarly for CD4M33, the assay started using 2 lg/ml peptide that, depending on the strain tested, was 3-to 10-fold higher than the IC 50 of CD4M33.…”