RESUMO
The interaction of the nucleocapsid NCp7 of the human immunodeficiency virus type 1 (HIV-1) Gag polyprotein with the RNA packaging signal Psi ensures specific encapsidation of the dimeric full length viral genome into nascent virus particles. Being an essential step in the HIV-1 replication cycle, specific genome encapsidation represents a promising target for therapeutic intervention. We previously selected peptides binding to HIV-1 Psi-RNA or stem loops (SL) thereof by phage display. Herein, we describe synthesis of peptide variants of the consensus HWWPWW motif on membrane supports to optimize Psi-RNA binding. The optimized peptide, psi-pepB, was characterized in detail with respect to its conformation and binding properties for the SL3 of the Psi packaging signal by NMR and tryptophan fluorescence quenching. Functional analysis revealed that psi-pepB caused a strong reduction of virus release by infected cells as monitored by reduced transduction efficiencies, capsid p24 antigen levels, and electron microscopy. Thus, this peptide shows antiviral activity and could serve as a lead compound to develop new drugs targeting HIV-1.
Assuntos
Fármacos Anti-HIV/farmacologia , Produtos do Gene gag/química , Genoma Viral , HIV-1/efeitos dos fármacos , Peptídeos/farmacologia , RNA Viral/química , Montagem de Vírus , Sequência de Aminoácidos , Linhagem Celular , HIV-1/genética , HIV-1/fisiologia , Humanos , Dados de Sequência MolecularRESUMO
Photo-CIDNP effects interpreted for individual residues are used for the structural characterization of non-native ensembles of proteins, which is described in this paper. Two-dimensional photo-CIDNP experiments are compared to conventional HSQC spectra to elucidate the relative solvent exposure of the six tryptophan residues in non-native states of hen egg white lysozyme. The differential solvent accessibility of the tryptophan residues in non-native lysozyme coincides with the dynamical properties of these residues monitored for both backbone and side chain NH sides obtained from analysis of transverse relaxation measurements. These data can be interpreted in the context of the hydrophobic clustering around the tryptophan residues and is supported by the application of this method to the cluster breaking W62G mutant of lysozyme.
