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Atomistic detailed mechanism and weak cation-conducting activity of HIV-1 Vpu revealed by free energy calculations.

Padhi S, Burri RR, Jameel S, Priyakumar UD - PLoS ONE (2014)

Bottom Line: Free energy profiles corresponding to the permeation of Na+ and K+ were found to be similar to each other indicating lack of ion selection, consistent with previous experimental studies.The Ser23 residue is shown to enhance ion transport via two mechanisms: creating a weak binding site, and increasing the effective hydrophilic length of the channel, both of which have previously been hypothesized in experiments.The results are consistent with previous conductance studies that showed Vpu to be a weakly conducting ion channel.

View Article: PubMed Central - PubMed

Affiliation: Centre for Computational Natural Sciences and Bioinformatics, International Institute of Information Technology, Hyderabad, India.

ABSTRACT
The viral protein U (Vpu) encoded by HIV-1 has been shown to assist in the detachment of virion particles from infected cells. Vpu forms cation-specific ion channels in host cells, and has been proposed as a potential drug target. An understanding of the mechanism of ion transport through Vpu is desirable, but remains limited because of the unavailability of an experimental structure of the channel. Using a structure of the pentameric form of Vpu--modeled and validated based on available experimental data--umbrella sampling molecular dynamics simulations (cumulative simulation time of more than 0.4 µs) were employed to elucidate the energetics and the molecular mechanism of ion transport in Vpu. Free energy profiles corresponding to the permeation of Na+ and K+ were found to be similar to each other indicating lack of ion selection, consistent with previous experimental studies. The Ser23 residue is shown to enhance ion transport via two mechanisms: creating a weak binding site, and increasing the effective hydrophilic length of the channel, both of which have previously been hypothesized in experiments. A two-dimensional free energy landscape has been computed to model multiple ion permeation, based on which a mechanism for ion conduction is proposed. It is shown that only one ion can pass through the channel at a time. This, along with a stretch of hydrophobic residues in the transmembrane domain of Vpu, explains the slow kinetics of ion conduction. The results are consistent with previous conductance studies that showed Vpu to be a weakly conducting ion channel.

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Electrostatic profile of pore-lining residues.(A) The pentamer model used in the study shown with the Ser23 residue in van der Waals representation. (B) A representation of the electrostatic surface of pore-lining residues in the channel. The monomer unit on the front has been omitted for clarity. Red indicates negatively charged regions, blue indicates positively charged regions, and white indicates nonpolar regions. Only the sidechain atoms are shown in color, and the backbone atoms are in white to allow a clear illustration of the nature of pore-lining residues.
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pone-0112983-g002: Electrostatic profile of pore-lining residues.(A) The pentamer model used in the study shown with the Ser23 residue in van der Waals representation. (B) A representation of the electrostatic surface of pore-lining residues in the channel. The monomer unit on the front has been omitted for clarity. Red indicates negatively charged regions, blue indicates positively charged regions, and white indicates nonpolar regions. Only the sidechain atoms are shown in color, and the backbone atoms are in white to allow a clear illustration of the nature of pore-lining residues.

Mentions: A representation of the channel modeled in a hydrated lipid bilayer is shown in Figure 1 (B). A few pore water molecules can be seen, especially in the bottom half of the pore, towards the C-terminal opening. The channel consists of five helical TM domains held together predominantly by van der Waals forces, with the interface between the helices being formed by nonpolar residues [24]. The channel is wider at the C-terminal end than at the N-terminal end, leading to a relatively greater degree of hydration at this end. The residues Arg30 and Tyr29 at the C-terminal end of the TM domain face the exterior side of the channel, and they stabilize the protein in the lipid bilayer by forming hydrogen bonds with phospholipid headgroups [24]. The sidechain of Arg30 of each of the monomers is also able to form a salt bridge with a Glu28 residue on the adjacent monomer. Although Lys31 is directed towards the interior of the channel, it lies in a region that is sufficiently solvated, so that the residue is shielded from exerting any influence on any ion passing through the channel. The orientation of these residues in the channel is shown in Figure S1 (A). A serine at position 23 faces the lumen of the pore, and it marks the end of the small hydrophilic space at the C-terminal entrance of the channel. Figure 2 (A) shows the serine residue facing the interior of the pentameric channel. The pore is constricted in the middle, which is a largely dehydrated region, owing to the occurrence of hydrophobic residues. The kinetics of ion conduction through the pore is likely to be controlled by this long hydrophobic stretch. Initially, the nature of the pore was investigated by examining the electrostatic representation of the pore-lining residues in the channel shown in Figure 2 (B). In the surface representation, one monomer of Vpu has been omitted in the figure for clarity. The figure shows electronegative regions in red, and electropositive regions in blue. It can be seen that the top and middle region of the channel is constricted and lined entirely by nonpolar residues. Further down the channel, towards the C-terminal end, the pore widens, and the occurrence of a serine residue gives rise to a slightly hydrophilic region. The widening of the pore was shown to be due to the kink around the Ile17 residue (see Figure S1 (B)) [24], which is in agreement with experimental data [19]. The serine residue can be seen as a red region at the bottom of the channel. The occurrence of such a hydrophilic region is expected to stabilize the permeating ion, thereby assisting ion transport. It follows that the length of the hydrophobic stretch would have been greater in the absence of this serine residue. The serine therefore reduces the effective length of the hydrophobic stretch, and notably, it has been reported previously that the Ser23→Leu mutant of Vpu is inactive towards ion permeation [28].


Atomistic detailed mechanism and weak cation-conducting activity of HIV-1 Vpu revealed by free energy calculations.

Padhi S, Burri RR, Jameel S, Priyakumar UD - PLoS ONE (2014)

Electrostatic profile of pore-lining residues.(A) The pentamer model used in the study shown with the Ser23 residue in van der Waals representation. (B) A representation of the electrostatic surface of pore-lining residues in the channel. The monomer unit on the front has been omitted for clarity. Red indicates negatively charged regions, blue indicates positively charged regions, and white indicates nonpolar regions. Only the sidechain atoms are shown in color, and the backbone atoms are in white to allow a clear illustration of the nature of pore-lining residues.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231112&req=5

pone-0112983-g002: Electrostatic profile of pore-lining residues.(A) The pentamer model used in the study shown with the Ser23 residue in van der Waals representation. (B) A representation of the electrostatic surface of pore-lining residues in the channel. The monomer unit on the front has been omitted for clarity. Red indicates negatively charged regions, blue indicates positively charged regions, and white indicates nonpolar regions. Only the sidechain atoms are shown in color, and the backbone atoms are in white to allow a clear illustration of the nature of pore-lining residues.
Mentions: A representation of the channel modeled in a hydrated lipid bilayer is shown in Figure 1 (B). A few pore water molecules can be seen, especially in the bottom half of the pore, towards the C-terminal opening. The channel consists of five helical TM domains held together predominantly by van der Waals forces, with the interface between the helices being formed by nonpolar residues [24]. The channel is wider at the C-terminal end than at the N-terminal end, leading to a relatively greater degree of hydration at this end. The residues Arg30 and Tyr29 at the C-terminal end of the TM domain face the exterior side of the channel, and they stabilize the protein in the lipid bilayer by forming hydrogen bonds with phospholipid headgroups [24]. The sidechain of Arg30 of each of the monomers is also able to form a salt bridge with a Glu28 residue on the adjacent monomer. Although Lys31 is directed towards the interior of the channel, it lies in a region that is sufficiently solvated, so that the residue is shielded from exerting any influence on any ion passing through the channel. The orientation of these residues in the channel is shown in Figure S1 (A). A serine at position 23 faces the lumen of the pore, and it marks the end of the small hydrophilic space at the C-terminal entrance of the channel. Figure 2 (A) shows the serine residue facing the interior of the pentameric channel. The pore is constricted in the middle, which is a largely dehydrated region, owing to the occurrence of hydrophobic residues. The kinetics of ion conduction through the pore is likely to be controlled by this long hydrophobic stretch. Initially, the nature of the pore was investigated by examining the electrostatic representation of the pore-lining residues in the channel shown in Figure 2 (B). In the surface representation, one monomer of Vpu has been omitted in the figure for clarity. The figure shows electronegative regions in red, and electropositive regions in blue. It can be seen that the top and middle region of the channel is constricted and lined entirely by nonpolar residues. Further down the channel, towards the C-terminal end, the pore widens, and the occurrence of a serine residue gives rise to a slightly hydrophilic region. The widening of the pore was shown to be due to the kink around the Ile17 residue (see Figure S1 (B)) [24], which is in agreement with experimental data [19]. The serine residue can be seen as a red region at the bottom of the channel. The occurrence of such a hydrophilic region is expected to stabilize the permeating ion, thereby assisting ion transport. It follows that the length of the hydrophobic stretch would have been greater in the absence of this serine residue. The serine therefore reduces the effective length of the hydrophobic stretch, and notably, it has been reported previously that the Ser23→Leu mutant of Vpu is inactive towards ion permeation [28].

Bottom Line: Free energy profiles corresponding to the permeation of Na+ and K+ were found to be similar to each other indicating lack of ion selection, consistent with previous experimental studies.The Ser23 residue is shown to enhance ion transport via two mechanisms: creating a weak binding site, and increasing the effective hydrophilic length of the channel, both of which have previously been hypothesized in experiments.The results are consistent with previous conductance studies that showed Vpu to be a weakly conducting ion channel.

View Article: PubMed Central - PubMed

Affiliation: Centre for Computational Natural Sciences and Bioinformatics, International Institute of Information Technology, Hyderabad, India.

ABSTRACT
The viral protein U (Vpu) encoded by HIV-1 has been shown to assist in the detachment of virion particles from infected cells. Vpu forms cation-specific ion channels in host cells, and has been proposed as a potential drug target. An understanding of the mechanism of ion transport through Vpu is desirable, but remains limited because of the unavailability of an experimental structure of the channel. Using a structure of the pentameric form of Vpu--modeled and validated based on available experimental data--umbrella sampling molecular dynamics simulations (cumulative simulation time of more than 0.4 µs) were employed to elucidate the energetics and the molecular mechanism of ion transport in Vpu. Free energy profiles corresponding to the permeation of Na+ and K+ were found to be similar to each other indicating lack of ion selection, consistent with previous experimental studies. The Ser23 residue is shown to enhance ion transport via two mechanisms: creating a weak binding site, and increasing the effective hydrophilic length of the channel, both of which have previously been hypothesized in experiments. A two-dimensional free energy landscape has been computed to model multiple ion permeation, based on which a mechanism for ion conduction is proposed. It is shown that only one ion can pass through the channel at a time. This, along with a stretch of hydrophobic residues in the transmembrane domain of Vpu, explains the slow kinetics of ion conduction. The results are consistent with previous conductance studies that showed Vpu to be a weakly conducting ion channel.

Show MeSH
Related in: MedlinePlus