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Behavior of solvent-exposed hydrophobic groove in the anti-apoptotic Bcl-XL protein: clues for its ability to bind diverse BH3 ligands from MD simulations.

Lama D, Modi V, Sankararamakrishnan R - PLoS ONE (2013)

Bottom Line: The solvent accessible surface areas of bulky hydrophobic residues in the groove are significantly buried by the loop LB connecting the helix H2 and subsequent helix.These observations help to understand how the hydrophobic patch in Bcl-XL remains stable in the solvent-exposed state.We suggest that both the destabilization of helix H2 and the conformational heterogeneity of loop LB are important factors for binding of diverse ligands in the hydrophobic groove of Bcl-XL.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences & Bioengineering, Indian Institute of Technology Kanpur, Kanpur, India.

ABSTRACT
Bcl-XL is a member of Bcl-2 family of proteins involved in the regulation of intrinsic pathway of apoptosis. Its overexpression in many human cancers makes it an important target for anti-cancer drugs. Bcl-XL interacts with the BH3 domain of several pro-apoptotic Bcl-2 partners. This helical bundle protein has a pronounced hydrophobic groove which acts as a binding region for the BH3 domains. Eight independent molecular dynamics simulations of the apo/holo forms of Bcl-XL were carried out to investigate the behavior of solvent-exposed hydrophobic groove. The simulations used either a twin-range cut-off or particle mesh Ewald (PME) scheme to treat long-range interactions. Destabilization of the BH3 domain-containing helix H2 was observed in all four twin-range cut-off simulations. Most of the other major helices remained stable. The unwinding of H2 can be related to the ability of Bcl-XL to bind diverse BH3 ligands. The loss of helical character can also be linked to the formation of homo- or hetero-dimers in Bcl-2 proteins. Several experimental studies have suggested that exposure of BH3 domain is a crucial event before they form dimers. Thus unwinding of H2 seems to be functionally very important. The four PME simulations, however, revealed a stable helix H2. It is possible that the H2 unfolding might occur in PME simulations at longer time scales. Hydrophobic residues in the hydrophobic groove are involved in stable interactions among themselves. The solvent accessible surface areas of bulky hydrophobic residues in the groove are significantly buried by the loop LB connecting the helix H2 and subsequent helix. These observations help to understand how the hydrophobic patch in Bcl-XL remains stable in the solvent-exposed state. We suggest that both the destabilization of helix H2 and the conformational heterogeneity of loop LB are important factors for binding of diverse ligands in the hydrophobic groove of Bcl-XL.

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Flexible loop LB in structures of different Bcl-XL complexes.Superimposition of 13 experimentally determined structures of Bcl-XL complexes (black) on the apo-Bcl-XL structure (PDB ID: 1PQ0 shown in red). Although more structures of complexes are available, we have chosen those complexes in which the ligands are unique. The Cα-atoms of helices H2, H3 and H4 of structures were superposed on the same helical segments of apo structure. This highlights the conformational heterogeneity of loop LB which connects H2 and H3. The PDB IDs of the structures of Bcl-XL complexes are 1BXL, 1G5J, 1PQ1, 1YSG, 1YSI, 2P1L, 2YJ1, 2YXJ, 3FDM, 3INQ, 3PL7, 3QKD and 3R85.
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pone-0054397-g007: Flexible loop LB in structures of different Bcl-XL complexes.Superimposition of 13 experimentally determined structures of Bcl-XL complexes (black) on the apo-Bcl-XL structure (PDB ID: 1PQ0 shown in red). Although more structures of complexes are available, we have chosen those complexes in which the ligands are unique. The Cα-atoms of helices H2, H3 and H4 of structures were superposed on the same helical segments of apo structure. This highlights the conformational heterogeneity of loop LB which connects H2 and H3. The PDB IDs of the structures of Bcl-XL complexes are 1BXL, 1G5J, 1PQ1, 1YSG, 1YSI, 2P1L, 2YJ1, 2YXJ, 3FDM, 3INQ, 3PL7, 3QKD and 3R85.

Mentions: The stability of exposed hydrophobic groove can be explained by two factors: (i) interactions among the hydrophobic residues in the groove and (ii) burial of hydrophobic amino acids by the residues from loop LB. Interactions between exposed hydrophobic residues is akin to hydrophobic collapse when a protein is on the way to its folding process trying to shield its hydrophobic residues from solvent exposure. Hydrophobic residues are also protected from the solvent by loop LB. This loop connects the BH3-containing helix H2 with the next helix H3 and it wraps around the hydrophobic groove (Figure 1). In the apo-Bcl-XL structure, a short stretch of this loop assumes helical conformation while in the Bcl-XL structures in complex with pro-apoptotic BH3 peptides, it essentially adopts a random conformation. It has been shown that the loop LB has stable interactions with the bound BH3 peptides [30]. The present simulation studies clearly demonstrate that this loop plays a major role in shielding the hydrophobic residues in the hydrophobic groove of Bcl-XL. The flexible and dynamic nature of this loop is evident by comparing the apo-Bcl-XL structure with the structures of other Bcl-2 members, structures of Bcl-XL complexes and also the mutant Bcl-XL structures [19], [21], [31], [37], [59], [60], [61]. We have identified 13 structures of Bcl-XL complexes in PDB in which the bound molecules are BH3 peptides or other organic ligands. Superposition of these structures (Figure 7) clearly demonstrates that loop LB displays conformational heterogeneity to accommodate and to have strong interactions with the ligands. In the absence of any bound ligands, this loop helps burying 250 to 380 Å2 surface area of bulky hydrophobic residues when all eight simulations are considered. The burial and exposure of loop LB residues is not uniform across all simulations. This indicates that loop LB is constantly sampling many conformations to optimally bury as many hydrophobic residues in the hydrophobic groove as possible. Thus loop LB plays a pivotal role in stabilizing the exposed hydrophobic residues in the hydrophobic groove in the absence of any bound ligand. The role of loop LB residues in conferring selectivity and higher affinities to different Bcl-XL ligands has to be explored further.


Behavior of solvent-exposed hydrophobic groove in the anti-apoptotic Bcl-XL protein: clues for its ability to bind diverse BH3 ligands from MD simulations.

Lama D, Modi V, Sankararamakrishnan R - PLoS ONE (2013)

Flexible loop LB in structures of different Bcl-XL complexes.Superimposition of 13 experimentally determined structures of Bcl-XL complexes (black) on the apo-Bcl-XL structure (PDB ID: 1PQ0 shown in red). Although more structures of complexes are available, we have chosen those complexes in which the ligands are unique. The Cα-atoms of helices H2, H3 and H4 of structures were superposed on the same helical segments of apo structure. This highlights the conformational heterogeneity of loop LB which connects H2 and H3. The PDB IDs of the structures of Bcl-XL complexes are 1BXL, 1G5J, 1PQ1, 1YSG, 1YSI, 2P1L, 2YJ1, 2YXJ, 3FDM, 3INQ, 3PL7, 3QKD and 3R85.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3585337&req=5

pone-0054397-g007: Flexible loop LB in structures of different Bcl-XL complexes.Superimposition of 13 experimentally determined structures of Bcl-XL complexes (black) on the apo-Bcl-XL structure (PDB ID: 1PQ0 shown in red). Although more structures of complexes are available, we have chosen those complexes in which the ligands are unique. The Cα-atoms of helices H2, H3 and H4 of structures were superposed on the same helical segments of apo structure. This highlights the conformational heterogeneity of loop LB which connects H2 and H3. The PDB IDs of the structures of Bcl-XL complexes are 1BXL, 1G5J, 1PQ1, 1YSG, 1YSI, 2P1L, 2YJ1, 2YXJ, 3FDM, 3INQ, 3PL7, 3QKD and 3R85.
Mentions: The stability of exposed hydrophobic groove can be explained by two factors: (i) interactions among the hydrophobic residues in the groove and (ii) burial of hydrophobic amino acids by the residues from loop LB. Interactions between exposed hydrophobic residues is akin to hydrophobic collapse when a protein is on the way to its folding process trying to shield its hydrophobic residues from solvent exposure. Hydrophobic residues are also protected from the solvent by loop LB. This loop connects the BH3-containing helix H2 with the next helix H3 and it wraps around the hydrophobic groove (Figure 1). In the apo-Bcl-XL structure, a short stretch of this loop assumes helical conformation while in the Bcl-XL structures in complex with pro-apoptotic BH3 peptides, it essentially adopts a random conformation. It has been shown that the loop LB has stable interactions with the bound BH3 peptides [30]. The present simulation studies clearly demonstrate that this loop plays a major role in shielding the hydrophobic residues in the hydrophobic groove of Bcl-XL. The flexible and dynamic nature of this loop is evident by comparing the apo-Bcl-XL structure with the structures of other Bcl-2 members, structures of Bcl-XL complexes and also the mutant Bcl-XL structures [19], [21], [31], [37], [59], [60], [61]. We have identified 13 structures of Bcl-XL complexes in PDB in which the bound molecules are BH3 peptides or other organic ligands. Superposition of these structures (Figure 7) clearly demonstrates that loop LB displays conformational heterogeneity to accommodate and to have strong interactions with the ligands. In the absence of any bound ligands, this loop helps burying 250 to 380 Å2 surface area of bulky hydrophobic residues when all eight simulations are considered. The burial and exposure of loop LB residues is not uniform across all simulations. This indicates that loop LB is constantly sampling many conformations to optimally bury as many hydrophobic residues in the hydrophobic groove as possible. Thus loop LB plays a pivotal role in stabilizing the exposed hydrophobic residues in the hydrophobic groove in the absence of any bound ligand. The role of loop LB residues in conferring selectivity and higher affinities to different Bcl-XL ligands has to be explored further.

Bottom Line: The solvent accessible surface areas of bulky hydrophobic residues in the groove are significantly buried by the loop LB connecting the helix H2 and subsequent helix.These observations help to understand how the hydrophobic patch in Bcl-XL remains stable in the solvent-exposed state.We suggest that both the destabilization of helix H2 and the conformational heterogeneity of loop LB are important factors for binding of diverse ligands in the hydrophobic groove of Bcl-XL.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences & Bioengineering, Indian Institute of Technology Kanpur, Kanpur, India.

ABSTRACT
Bcl-XL is a member of Bcl-2 family of proteins involved in the regulation of intrinsic pathway of apoptosis. Its overexpression in many human cancers makes it an important target for anti-cancer drugs. Bcl-XL interacts with the BH3 domain of several pro-apoptotic Bcl-2 partners. This helical bundle protein has a pronounced hydrophobic groove which acts as a binding region for the BH3 domains. Eight independent molecular dynamics simulations of the apo/holo forms of Bcl-XL were carried out to investigate the behavior of solvent-exposed hydrophobic groove. The simulations used either a twin-range cut-off or particle mesh Ewald (PME) scheme to treat long-range interactions. Destabilization of the BH3 domain-containing helix H2 was observed in all four twin-range cut-off simulations. Most of the other major helices remained stable. The unwinding of H2 can be related to the ability of Bcl-XL to bind diverse BH3 ligands. The loss of helical character can also be linked to the formation of homo- or hetero-dimers in Bcl-2 proteins. Several experimental studies have suggested that exposure of BH3 domain is a crucial event before they form dimers. Thus unwinding of H2 seems to be functionally very important. The four PME simulations, however, revealed a stable helix H2. It is possible that the H2 unfolding might occur in PME simulations at longer time scales. Hydrophobic residues in the hydrophobic groove are involved in stable interactions among themselves. The solvent accessible surface areas of bulky hydrophobic residues in the groove are significantly buried by the loop LB connecting the helix H2 and subsequent helix. These observations help to understand how the hydrophobic patch in Bcl-XL remains stable in the solvent-exposed state. We suggest that both the destabilization of helix H2 and the conformational heterogeneity of loop LB are important factors for binding of diverse ligands in the hydrophobic groove of Bcl-XL.

Show MeSH
Related in: MedlinePlus