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Minimum free energy path of ligand-induced transition in adenylate kinase.

Matsunaga Y, Fujisaki H, Terada T, Furuta T, Moritsugu K, Kidera A - PLoS Comput. Biol. (2012)

Bottom Line: It was found that the LID domain was able to partially close without the ligand, while the closure of the AMPbd domain required the ligand binding.It was also found that complete closure of the LID domain required the dehydration of solvents around the P-loop.These findings suggest that the interplay of the two different types of domain motion is an essential feature in the conformational transition of the enzyme.

View Article: PubMed Central - PubMed

Affiliation: Computational Science Research Program, RIKEN, Wako, Japan. ymatsunaga@riken.jp

ABSTRACT
Large-scale conformational changes in proteins involve barrier-crossing transitions on the complex free energy surfaces of high-dimensional space. Such rare events cannot be efficiently captured by conventional molecular dynamics simulations. Here we show that, by combining the on-the-fly string method and the multi-state Bennett acceptance ratio (MBAR) method, the free energy profile of a conformational transition pathway in Escherichia coli adenylate kinase can be characterized in a high-dimensional space. The minimum free energy paths of the conformational transitions in adenylate kinase were explored by the on-the-fly string method in 20-dimensional space spanned by the 20 largest-amplitude principal modes, and the free energy and various kinds of average physical quantities along the pathways were successfully evaluated by the MBAR method. The influence of ligand binding on the pathways was characterized in terms of rigid-body motions of the lid-shaped ATP-binding domain (LID) and the AMP-binding (AMPbd) domains. It was found that the LID domain was able to partially close without the ligand, while the closure of the AMPbd domain required the ligand binding. The transition state ensemble of the ligand bound form was identified as those structures characterized by highly specific binding of the ligand to the AMPbd domain, and was validated by unrestrained MD simulations. It was also found that complete closure of the LID domain required the dehydration of solvents around the P-loop. These findings suggest that the interplay of the two different types of domain motion is an essential feature in the conformational transition of the enzyme.

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Dehydration of an occluded water around the P-loop.The isosurface representation of the 3D distribution function of water oxygen (red) and hydrogen (white) around the P-loop at (A) , (B) 41, and (C) 42. The surfaces show the areas in which the atoms are distributed four times as probably as in the bulk phase. For comparison, the oxygens of crystal waters are shown for (D) the open (PDBid: 4ake) and (E) closed conformations (PDBid: 1ake). An occluded water molecule at  and the corresponding crystal water of the open form are indicated by the circles. (F) Two-dimensional PMF surface as a function of the MFEP images and the distances of the LID-CORE domains. The PMF is represented by the colored contour lines. Regions of physical events (AMP-binding and dehydration) are encircled.
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pcbi-1002555-g006: Dehydration of an occluded water around the P-loop.The isosurface representation of the 3D distribution function of water oxygen (red) and hydrogen (white) around the P-loop at (A) , (B) 41, and (C) 42. The surfaces show the areas in which the atoms are distributed four times as probably as in the bulk phase. For comparison, the oxygens of crystal waters are shown for (D) the open (PDBid: 4ake) and (E) closed conformations (PDBid: 1ake). An occluded water molecule at and the corresponding crystal water of the open form are indicated by the circles. (F) Two-dimensional PMF surface as a function of the MFEP images and the distances of the LID-CORE domains. The PMF is represented by the colored contour lines. Regions of physical events (AMP-binding and dehydration) are encircled.

Mentions: A remaining question is how closure of the LID domain follows that of the AMPbd domain. Unlike the AMP-binding pocket, the ATP-binding sites, including the P-loop, are surrounded by charged residues, which attract interfacial water molecules. Upon LID closure, most of these water molecules will be dehydrated from the enzyme, but some may remain occluded. To characterize the behaviors of these water molecules, the 3D distribution function of their oxygen and hydrogen constituents were calculated along the MFEP using the MBAR method (see Materials and Methods). Figures 6A, 6B, and 6C display the isosurface representations of the 3D distribution functions around the P-loop at , 41, and 42, respectively. The surfaces show the areas in which the atoms are distributed four times as probably as in the bulk phase. At , the ATP phosphates are not yet bound to the P-loop because an occluded water molecule (encircled) is wedged between the phosphate and the P-loop, inhibiting binding of ATP and and bending of the side-chain of “invariant lysine” (Lys13), a residue that plays a critical role in orienting the phosphates to the proper catalytic position [36]. This occluded water molecule may correspond to that found in the crystal structure of apo-AK (PDBid: 4ake) (Figure 6D, encircled). Figures 6B and 6C clearly demonstrate that, upon removal of this water molecule, the ATP phosphates begin binding to the P-loop. These observations were confirmed by plots of the PMF surface mapped onto a space defined by the LID-CORE distance versus the index of image (Figure 6F), which shows that the PMF decreases discontinuously upon dehydration followed by LID domain closure. Interestingly, compared with the crystal structure (PDBid: 1ake)(Figure 6E), the position of the ATP moiety is shifted to the AMP side by one monophosphate unit. This may be a consequence of early binding of the AMP moiety. At a later stage (around ), this mismatch was corrected to form the same binding mode as observed in the crystal structure. This reformation of the binding mode may be induced by the tight binding of ATP adenine to the LID-CORE domains, and will not occur in the real enzymatic system containing ATP and AMP instead of the bisubstrate analog inhibitor, Ap5A.


Minimum free energy path of ligand-induced transition in adenylate kinase.

Matsunaga Y, Fujisaki H, Terada T, Furuta T, Moritsugu K, Kidera A - PLoS Comput. Biol. (2012)

Dehydration of an occluded water around the P-loop.The isosurface representation of the 3D distribution function of water oxygen (red) and hydrogen (white) around the P-loop at (A) , (B) 41, and (C) 42. The surfaces show the areas in which the atoms are distributed four times as probably as in the bulk phase. For comparison, the oxygens of crystal waters are shown for (D) the open (PDBid: 4ake) and (E) closed conformations (PDBid: 1ake). An occluded water molecule at  and the corresponding crystal water of the open form are indicated by the circles. (F) Two-dimensional PMF surface as a function of the MFEP images and the distances of the LID-CORE domains. The PMF is represented by the colored contour lines. Regions of physical events (AMP-binding and dehydration) are encircled.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3369945&req=5

pcbi-1002555-g006: Dehydration of an occluded water around the P-loop.The isosurface representation of the 3D distribution function of water oxygen (red) and hydrogen (white) around the P-loop at (A) , (B) 41, and (C) 42. The surfaces show the areas in which the atoms are distributed four times as probably as in the bulk phase. For comparison, the oxygens of crystal waters are shown for (D) the open (PDBid: 4ake) and (E) closed conformations (PDBid: 1ake). An occluded water molecule at and the corresponding crystal water of the open form are indicated by the circles. (F) Two-dimensional PMF surface as a function of the MFEP images and the distances of the LID-CORE domains. The PMF is represented by the colored contour lines. Regions of physical events (AMP-binding and dehydration) are encircled.
Mentions: A remaining question is how closure of the LID domain follows that of the AMPbd domain. Unlike the AMP-binding pocket, the ATP-binding sites, including the P-loop, are surrounded by charged residues, which attract interfacial water molecules. Upon LID closure, most of these water molecules will be dehydrated from the enzyme, but some may remain occluded. To characterize the behaviors of these water molecules, the 3D distribution function of their oxygen and hydrogen constituents were calculated along the MFEP using the MBAR method (see Materials and Methods). Figures 6A, 6B, and 6C display the isosurface representations of the 3D distribution functions around the P-loop at , 41, and 42, respectively. The surfaces show the areas in which the atoms are distributed four times as probably as in the bulk phase. At , the ATP phosphates are not yet bound to the P-loop because an occluded water molecule (encircled) is wedged between the phosphate and the P-loop, inhibiting binding of ATP and and bending of the side-chain of “invariant lysine” (Lys13), a residue that plays a critical role in orienting the phosphates to the proper catalytic position [36]. This occluded water molecule may correspond to that found in the crystal structure of apo-AK (PDBid: 4ake) (Figure 6D, encircled). Figures 6B and 6C clearly demonstrate that, upon removal of this water molecule, the ATP phosphates begin binding to the P-loop. These observations were confirmed by plots of the PMF surface mapped onto a space defined by the LID-CORE distance versus the index of image (Figure 6F), which shows that the PMF decreases discontinuously upon dehydration followed by LID domain closure. Interestingly, compared with the crystal structure (PDBid: 1ake)(Figure 6E), the position of the ATP moiety is shifted to the AMP side by one monophosphate unit. This may be a consequence of early binding of the AMP moiety. At a later stage (around ), this mismatch was corrected to form the same binding mode as observed in the crystal structure. This reformation of the binding mode may be induced by the tight binding of ATP adenine to the LID-CORE domains, and will not occur in the real enzymatic system containing ATP and AMP instead of the bisubstrate analog inhibitor, Ap5A.

Bottom Line: It was found that the LID domain was able to partially close without the ligand, while the closure of the AMPbd domain required the ligand binding.It was also found that complete closure of the LID domain required the dehydration of solvents around the P-loop.These findings suggest that the interplay of the two different types of domain motion is an essential feature in the conformational transition of the enzyme.

View Article: PubMed Central - PubMed

Affiliation: Computational Science Research Program, RIKEN, Wako, Japan. ymatsunaga@riken.jp

ABSTRACT
Large-scale conformational changes in proteins involve barrier-crossing transitions on the complex free energy surfaces of high-dimensional space. Such rare events cannot be efficiently captured by conventional molecular dynamics simulations. Here we show that, by combining the on-the-fly string method and the multi-state Bennett acceptance ratio (MBAR) method, the free energy profile of a conformational transition pathway in Escherichia coli adenylate kinase can be characterized in a high-dimensional space. The minimum free energy paths of the conformational transitions in adenylate kinase were explored by the on-the-fly string method in 20-dimensional space spanned by the 20 largest-amplitude principal modes, and the free energy and various kinds of average physical quantities along the pathways were successfully evaluated by the MBAR method. The influence of ligand binding on the pathways was characterized in terms of rigid-body motions of the lid-shaped ATP-binding domain (LID) and the AMP-binding (AMPbd) domains. It was found that the LID domain was able to partially close without the ligand, while the closure of the AMPbd domain required the ligand binding. The transition state ensemble of the ligand bound form was identified as those structures characterized by highly specific binding of the ligand to the AMPbd domain, and was validated by unrestrained MD simulations. It was also found that complete closure of the LID domain required the dehydration of solvents around the P-loop. These findings suggest that the interplay of the two different types of domain motion is an essential feature in the conformational transition of the enzyme.

Show MeSH
Related in: MedlinePlus