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Equilibrated atomic models of outward-facing P-glycoprotein and effect of ATP binding on structural dynamics.

Pan L, Aller SG - Sci Rep (2015)

Bottom Line: Three long lasting (>100 ns) meta-stable states were apparent in the presence of MgATP revealing new insights into alternating access.The two ATP-binding pockets are highly asymmetric resulting in differential control of overall structural dynamics and allosteric regulation of the drug-binding pocket.Equilibrated Pgp has a considerably different electrostatic profile compared to Sav1866 that implicates significant kinetic and thermodynamic differences in transport mechanisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Toxicology, University of Alabama at Birmingham, 1025 18th Street South, Birmingham, AL, 35205 USA.

ABSTRACT
P-glycoprotein (Pgp) is an ATP-binding cassette (ABC) transporter that alternates between inward- and outward-facing conformations to capture and force substrates out of cells like a peristaltic pump. The high degree of similarity in outward-facing structures across evolution of ABC transporters allowed construction of a high-confidence outward-facing Pgp atomic model based on crystal structures of outward-facing Sav1866 and inward-facing Pgp. The model adhered to previous experimentally determined secondary- and tertiary- configurations during all-atom molecular dynamics simulations in the presence or absence of MgATP. Three long lasting (>100 ns) meta-stable states were apparent in the presence of MgATP revealing new insights into alternating access. The two ATP-binding pockets are highly asymmetric resulting in differential control of overall structural dynamics and allosteric regulation of the drug-binding pocket. Equilibrated Pgp has a considerably different electrostatic profile compared to Sav1866 that implicates significant kinetic and thermodynamic differences in transport mechanisms.

No MeSH data available.


Related in: MedlinePlus

Conformational change illustrated with landmark residues.Landmark residue distances at each regions of the protein are measured using the average of the last 50 ns trajectories from each simulation. The protein is shown as cartoon representation with half1 in pink and half2 in ice blue. The Cα of the landmark residues are drawn as spheres in green (D-L). P atoms of lipid bi-layer are drawn in semi-transparent sphere in gold. MgATP is drawn in licorice with H in white, N in blue, C in cyan, O in red, P in gold and Mg2+ in pink. Orange arrows point to the directions of conformational change. The conformational changes are monitored using residue pair from two pseudo-symmetric counterparts: Extracellular gate (A) uses A79-T736 (ECL1-ECL3), G207-G850(TM3-TM9), G325-E968(ECL2-ECL4), drug binding pocket (B) uses higher end F71(TM1)-F728(TM7) pair and lower end Q343 (TM6)-Q986 (TM12) pair, ATP-binding sites (C) uses K429-S1173 (site 1) and K1072-S528 (site 2). The predominant conformation observed from all three runs of apo-Pgp (E, G, I and K) and predominant conformation observed from 30 ns-100 ns of run1, 30-50 ns of run2 and 30-200 ns of run3 of Pgp-MgATP (F, H, J and L) are different in various aspects.
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f6: Conformational change illustrated with landmark residues.Landmark residue distances at each regions of the protein are measured using the average of the last 50 ns trajectories from each simulation. The protein is shown as cartoon representation with half1 in pink and half2 in ice blue. The Cα of the landmark residues are drawn as spheres in green (D-L). P atoms of lipid bi-layer are drawn in semi-transparent sphere in gold. MgATP is drawn in licorice with H in white, N in blue, C in cyan, O in red, P in gold and Mg2+ in pink. Orange arrows point to the directions of conformational change. The conformational changes are monitored using residue pair from two pseudo-symmetric counterparts: Extracellular gate (A) uses A79-T736 (ECL1-ECL3), G207-G850(TM3-TM9), G325-E968(ECL2-ECL4), drug binding pocket (B) uses higher end F71(TM1)-F728(TM7) pair and lower end Q343 (TM6)-Q986 (TM12) pair, ATP-binding sites (C) uses K429-S1173 (site 1) and K1072-S528 (site 2). The predominant conformation observed from all three runs of apo-Pgp (E, G, I and K) and predominant conformation observed from 30 ns-100 ns of run1, 30-50 ns of run2 and 30-200 ns of run3 of Pgp-MgATP (F, H, J and L) are different in various aspects.

Mentions: The conformational changes were monitored using residue pairs from pseudo-symmetric counterparts (Figure 6). Six different averaged pair distances at the “extracellular gate” in the six simulations demonstrated large randomness in this region (Figure 6A). Therefore, large flexibility and numerous meta-stable states with similar energy are implicated. In addition, it was also shown that the three residue pairs are strongly inter-correlated in distance patterns, which suggests that the opening/closing of the extracellular gate occurs simultaneously for all TMs. When compared to the initial structure, the A79-T736 pair distance markedly decreased for all simulations whereas the other pairs at the extracellular gate fluctuated within a small dynamic range from the initial distance (Figure 6 A, G and H). The simulations demonstrate that the dynamic structure of the extracellular gate oscillates between open and closed forms rather than a relatively fixed conformation.


Equilibrated atomic models of outward-facing P-glycoprotein and effect of ATP binding on structural dynamics.

Pan L, Aller SG - Sci Rep (2015)

Conformational change illustrated with landmark residues.Landmark residue distances at each regions of the protein are measured using the average of the last 50 ns trajectories from each simulation. The protein is shown as cartoon representation with half1 in pink and half2 in ice blue. The Cα of the landmark residues are drawn as spheres in green (D-L). P atoms of lipid bi-layer are drawn in semi-transparent sphere in gold. MgATP is drawn in licorice with H in white, N in blue, C in cyan, O in red, P in gold and Mg2+ in pink. Orange arrows point to the directions of conformational change. The conformational changes are monitored using residue pair from two pseudo-symmetric counterparts: Extracellular gate (A) uses A79-T736 (ECL1-ECL3), G207-G850(TM3-TM9), G325-E968(ECL2-ECL4), drug binding pocket (B) uses higher end F71(TM1)-F728(TM7) pair and lower end Q343 (TM6)-Q986 (TM12) pair, ATP-binding sites (C) uses K429-S1173 (site 1) and K1072-S528 (site 2). The predominant conformation observed from all three runs of apo-Pgp (E, G, I and K) and predominant conformation observed from 30 ns-100 ns of run1, 30-50 ns of run2 and 30-200 ns of run3 of Pgp-MgATP (F, H, J and L) are different in various aspects.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4389535&req=5

f6: Conformational change illustrated with landmark residues.Landmark residue distances at each regions of the protein are measured using the average of the last 50 ns trajectories from each simulation. The protein is shown as cartoon representation with half1 in pink and half2 in ice blue. The Cα of the landmark residues are drawn as spheres in green (D-L). P atoms of lipid bi-layer are drawn in semi-transparent sphere in gold. MgATP is drawn in licorice with H in white, N in blue, C in cyan, O in red, P in gold and Mg2+ in pink. Orange arrows point to the directions of conformational change. The conformational changes are monitored using residue pair from two pseudo-symmetric counterparts: Extracellular gate (A) uses A79-T736 (ECL1-ECL3), G207-G850(TM3-TM9), G325-E968(ECL2-ECL4), drug binding pocket (B) uses higher end F71(TM1)-F728(TM7) pair and lower end Q343 (TM6)-Q986 (TM12) pair, ATP-binding sites (C) uses K429-S1173 (site 1) and K1072-S528 (site 2). The predominant conformation observed from all three runs of apo-Pgp (E, G, I and K) and predominant conformation observed from 30 ns-100 ns of run1, 30-50 ns of run2 and 30-200 ns of run3 of Pgp-MgATP (F, H, J and L) are different in various aspects.
Mentions: The conformational changes were monitored using residue pairs from pseudo-symmetric counterparts (Figure 6). Six different averaged pair distances at the “extracellular gate” in the six simulations demonstrated large randomness in this region (Figure 6A). Therefore, large flexibility and numerous meta-stable states with similar energy are implicated. In addition, it was also shown that the three residue pairs are strongly inter-correlated in distance patterns, which suggests that the opening/closing of the extracellular gate occurs simultaneously for all TMs. When compared to the initial structure, the A79-T736 pair distance markedly decreased for all simulations whereas the other pairs at the extracellular gate fluctuated within a small dynamic range from the initial distance (Figure 6 A, G and H). The simulations demonstrate that the dynamic structure of the extracellular gate oscillates between open and closed forms rather than a relatively fixed conformation.

Bottom Line: Three long lasting (>100 ns) meta-stable states were apparent in the presence of MgATP revealing new insights into alternating access.The two ATP-binding pockets are highly asymmetric resulting in differential control of overall structural dynamics and allosteric regulation of the drug-binding pocket.Equilibrated Pgp has a considerably different electrostatic profile compared to Sav1866 that implicates significant kinetic and thermodynamic differences in transport mechanisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Toxicology, University of Alabama at Birmingham, 1025 18th Street South, Birmingham, AL, 35205 USA.

ABSTRACT
P-glycoprotein (Pgp) is an ATP-binding cassette (ABC) transporter that alternates between inward- and outward-facing conformations to capture and force substrates out of cells like a peristaltic pump. The high degree of similarity in outward-facing structures across evolution of ABC transporters allowed construction of a high-confidence outward-facing Pgp atomic model based on crystal structures of outward-facing Sav1866 and inward-facing Pgp. The model adhered to previous experimentally determined secondary- and tertiary- configurations during all-atom molecular dynamics simulations in the presence or absence of MgATP. Three long lasting (>100 ns) meta-stable states were apparent in the presence of MgATP revealing new insights into alternating access. The two ATP-binding pockets are highly asymmetric resulting in differential control of overall structural dynamics and allosteric regulation of the drug-binding pocket. Equilibrated Pgp has a considerably different electrostatic profile compared to Sav1866 that implicates significant kinetic and thermodynamic differences in transport mechanisms.

No MeSH data available.


Related in: MedlinePlus