NMR identification of calcineurin B residues affected by binding of a calcineurin A peptide

AbstractTriple resonance 3D NMR methods have been used to study the interaction between calcineurin B and a peptide fragment of calcineurin A for which it has high affinity (KD ∼4 × 10−7 M). Although calcineurin B aggregates at NMR concentrations of ∼ 1 mM, in the presence of a target peptide fragment of calcineurin A it becomes monomeric and yields NMR spectra that are very similar to those reported previously for calcineurin B solubilized by the zwitterionic detergent CHAPS. Changes in chemical shifts between CHAPS- and peptide-solubilized calcineurin B are small which is indicative of no differences in secondary structure. Residues most affected by binding to target peptide are found primarily on the hydrophobic faces of the four helices, present in each of the two globular domains in calcineurin B, and in the loops connecting helices II and III, IV and V, and possibly in the C-terminal 12 residues, which also exhibit a change in mobility

Alternative Conformations of HIV-1 V3 Loops Mimic β Hairpins in Chemokines, Suggesting a Mechanism for Coreceptor Selectivity

AbstractThe V3 loop of the HIV-1 envelope glycoprotein gp120 is involved in binding to the CCR5 and CXCR4 coreceptors. The structure of an HIV-1MN V3 peptide bound to the Fv of the broadly neutralizing human monoclonal antibody 447-52D was solved by NMR and found to be a β hairpin. This structure of V3MN was found to have conformation and sequence similarities to β hairpins in CD8 and CCR5 ligands MIP-1α, MIP-1β, and RANTES and differed from the β hairpin of a V3IIIB peptide bound to the strain-specific murine anti-gp120IIIB antibody 0.5β. In contrast to the structure of the bound V3MN peptide, the V3IIIB peptide resembles a β hairpin in SDF-1, a CXCR4 ligand. These data suggest that the 447-52D-bound V3MN and the 0.5β-bound V3IIIB structures represent alternative V3 conformations responsible for selective interactions with CCR5 and CXCR4, respectively

The principal neutralizing determinant of HIV-1 located in V3 of gp120 forms a 12-residue loop by internal hydrophobic interactions

AbstractThe interactions of the peptide RP135a (RKSIRIQRGPGRAFVT), corresponding to residues 311–326 of gp120 of HIV-1IIIB, with the anti-gp120 HIV-1IIIB neutralizing antibody 0.5β were studied by NMR. The NOESY difference spectra measured using specifically deuterated derivatives of the peptide show exclusively the interactions of the deuterated residues both within the bound peptide and with the Fab fragment of the antibody. These measurements reveal hydrophobic interactions within the bound peptide between Ile-4, Ile-6 and Val-15 that create a 12-residue loop with these residues at the base and the conserved GPGR sequence at its top