RESUMO
Part of the genome of the human immunodeficiency virus type 1 (HIV-1) encodes for a short membrane protein Vpu, which has a length of 81 amino acids. It has two functional roles: (i) to downregulate CD4 and (ii) to support particle release. These roles are attributed to two distinct domains of the peptide, the cytoplasmic and transmembrane (TM) domains, respectively. It has been suggested that the enhanced particle release function is linked to the ion channel activity of Vpu, with a slight preference for cations over anions. To allow ion flux across the membrane Vpu would be required to assemble in homooligomers to form functional water-filled pores. In this study molecular dynamics simulations are used to address the role of particular amino acids in 4, 5, and 6 TM helix bundle structures. The helices (Vpu(6-33)) are extended to include hydrophilic residues such as Glu, Tyr, and Arg (EYR motif). Our simulations indicate that this motif destabilizes the bundles at their C-terminal ends. The arginines point into the pore to form a positive charged ring that could act as a putative selectivity filter. The helices of the bundles adopt slightly higher average tilt angles with decreasing number of helices. We also suggest that the helices are kinked. Conductance measurements on a peptide (Vpu(1-32)) reconstituted into lipid membranes show that the peptide forms ion channels with several conductance levels.
Assuntos
Simulação por Computador , HIV-1/química , Proteínas de Membrana/química , Modelos Moleculares , Fragmentos de Peptídeos/química , Proteínas Virais Reguladoras e Acessórias/química , Sequência de Aminoácidos , Condutividade Elétrica , HIV-1/fisiologia , Proteínas do Vírus da Imunodeficiência Humana , Humanos , Potenciais da Membrana , Proteínas de Membrana/fisiologia , Dados de Sequência Molecular , Técnicas de Patch-Clamp , Fragmentos de Peptídeos/fisiologia , Conformação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Proteínas Virais Reguladoras e Acessórias/fisiologia , Água/químicaRESUMO
Vpu is an 81 amino acid auxiliary protein in HIV-1 which exhibits channel activity. We used two homo-pentameric bundles with the helical transmembrane segments derived from FTIR spectroscopy in combination with a global molecular dynamics search protocol: (i) tryptophans (W) pointing into the pore, and (ii) W facing the lipids. Two equivalent bundles have been generated using a simulated annealing via a restrained molecular dynamics simulations (SA/MD) protocol. A fifth model was generated via SA/MD with all serines facing the pore. The latter model adopts a very stable structure during the 2 ns of simulation. The stability of the models with W facing the pore depends on the starting structure. A possible gating mechanism is outlined.