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Jill E. Gready - One of the best experts on this subject based on the ideXlab platform.
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Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr): Protonation State of the ligand and active-site residues.
The Journal of Physical Chemistry B, 2009Co-Authors: Hernan Alonso, Peter L. Cummins, Jill E. GreadyAbstract:Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) catalyzes the transfer of the N5-methyl group from N5-methyltetrahydrofolate (CH(3)THF) to the cobalt center of a corrinoid/iron-sulfur protein, a reaction similar to that of cobalamin-dependent methionine synthase (MetH). For such a reaction to occur, CH(3)THF is expected to be activated by a stereospecific Protonation at the N5 position. It has been shown experimentally that binding to MeTr is associated with a pK(a) increase and proton uptake. The enzyme could achieve this by binding the unprotonated form of CH(3)THF, followed by specific Protonation at the correct orientation. Here we have used computational approaches to investigate the Protonation State of the ligand and active-site residues in MeTr. First, quantum mechanical (QM) methods with the PCM solvation model were used to predict Protonation positions and pK(a) values of pterin, folate, and their analogues in an aqueous environment. After a reliable calibration of computational and experimental results was obtained, the effect of the protein environment was then considered. Different Protonation States of CH(3)THF and active-site aspartic residues (D75 and D160) were investigated using QM calculations of active-site fragment complexes and the perturbed quantum atom (PQA) approach within QM/MM simulations. The final free energy results indicate that the N5 position of the tetrahydropterin ring is the preferred Protonation position of CH(3)THF when bound to the active site of MeTr, followed by Asp160. We also found that the active-site environment is likely to increase the pK(a) of N5 by about 3 units, leading to proton uptake upon CH(3)THF binding, as observed experimentally for MeTr. Some implications of the results are discussed for the MetH mechanism.
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methyltetrahydrofolate corrinoid iron sulfur protein methyltransferase metr Protonation State of the ligand and active site residues
Journal of Physical Chemistry B, 2009Co-Authors: Hernan Alonso, Peter L. Cummins, Jill E. GreadyAbstract:Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) catalyzes the transfer of the N5-methyl group from N5-methyltetrahydrofolate (CH(3)THF) to the cobalt center of a corrinoid/iron-sulfur protein, a reaction similar to that of cobalamin-dependent methionine synthase (MetH). For such a reaction to occur, CH(3)THF is expected to be activated by a stereospecific Protonation at the N5 position. It has been shown experimentally that binding to MeTr is associated with a pK(a) increase and proton uptake. The enzyme could achieve this by binding the unprotonated form of CH(3)THF, followed by specific Protonation at the correct orientation. Here we have used computational approaches to investigate the Protonation State of the ligand and active-site residues in MeTr. First, quantum mechanical (QM) methods with the PCM solvation model were used to predict Protonation positions and pK(a) values of pterin, folate, and their analogues in an aqueous environment. After a reliable calibration of computational and experimental results was obtained, the effect of the protein environment was then considered. Different Protonation States of CH(3)THF and active-site aspartic residues (D75 and D160) were investigated using QM calculations of active-site fragment complexes and the perturbed quantum atom (PQA) approach within QM/MM simulations. The final free energy results indicate that the N5 position of the tetrahydropterin ring is the preferred Protonation position of CH(3)THF when bound to the active site of MeTr, followed by Asp160. We also found that the active-site environment is likely to increase the pK(a) of N5 by about 3 units, leading to proton uptake upon CH(3)THF binding, as observed experimentally for MeTr. Some implications of the results are discussed for the MetH mechanism.
Hernan Alonso - One of the best experts on this subject based on the ideXlab platform.
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Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr): Protonation State of the ligand and active-site residues.
The Journal of Physical Chemistry B, 2009Co-Authors: Hernan Alonso, Peter L. Cummins, Jill E. GreadyAbstract:Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) catalyzes the transfer of the N5-methyl group from N5-methyltetrahydrofolate (CH(3)THF) to the cobalt center of a corrinoid/iron-sulfur protein, a reaction similar to that of cobalamin-dependent methionine synthase (MetH). For such a reaction to occur, CH(3)THF is expected to be activated by a stereospecific Protonation at the N5 position. It has been shown experimentally that binding to MeTr is associated with a pK(a) increase and proton uptake. The enzyme could achieve this by binding the unprotonated form of CH(3)THF, followed by specific Protonation at the correct orientation. Here we have used computational approaches to investigate the Protonation State of the ligand and active-site residues in MeTr. First, quantum mechanical (QM) methods with the PCM solvation model were used to predict Protonation positions and pK(a) values of pterin, folate, and their analogues in an aqueous environment. After a reliable calibration of computational and experimental results was obtained, the effect of the protein environment was then considered. Different Protonation States of CH(3)THF and active-site aspartic residues (D75 and D160) were investigated using QM calculations of active-site fragment complexes and the perturbed quantum atom (PQA) approach within QM/MM simulations. The final free energy results indicate that the N5 position of the tetrahydropterin ring is the preferred Protonation position of CH(3)THF when bound to the active site of MeTr, followed by Asp160. We also found that the active-site environment is likely to increase the pK(a) of N5 by about 3 units, leading to proton uptake upon CH(3)THF binding, as observed experimentally for MeTr. Some implications of the results are discussed for the MetH mechanism.
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methyltetrahydrofolate corrinoid iron sulfur protein methyltransferase metr Protonation State of the ligand and active site residues
Journal of Physical Chemistry B, 2009Co-Authors: Hernan Alonso, Peter L. Cummins, Jill E. GreadyAbstract:Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) catalyzes the transfer of the N5-methyl group from N5-methyltetrahydrofolate (CH(3)THF) to the cobalt center of a corrinoid/iron-sulfur protein, a reaction similar to that of cobalamin-dependent methionine synthase (MetH). For such a reaction to occur, CH(3)THF is expected to be activated by a stereospecific Protonation at the N5 position. It has been shown experimentally that binding to MeTr is associated with a pK(a) increase and proton uptake. The enzyme could achieve this by binding the unprotonated form of CH(3)THF, followed by specific Protonation at the correct orientation. Here we have used computational approaches to investigate the Protonation State of the ligand and active-site residues in MeTr. First, quantum mechanical (QM) methods with the PCM solvation model were used to predict Protonation positions and pK(a) values of pterin, folate, and their analogues in an aqueous environment. After a reliable calibration of computational and experimental results was obtained, the effect of the protein environment was then considered. Different Protonation States of CH(3)THF and active-site aspartic residues (D75 and D160) were investigated using QM calculations of active-site fragment complexes and the perturbed quantum atom (PQA) approach within QM/MM simulations. The final free energy results indicate that the N5 position of the tetrahydropterin ring is the preferred Protonation position of CH(3)THF when bound to the active site of MeTr, followed by Asp160. We also found that the active-site environment is likely to increase the pK(a) of N5 by about 3 units, leading to proton uptake upon CH(3)THF binding, as observed experimentally for MeTr. Some implications of the results are discussed for the MetH mechanism.
Peter L. Cummins - One of the best experts on this subject based on the ideXlab platform.
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Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr): Protonation State of the ligand and active-site residues.
The Journal of Physical Chemistry B, 2009Co-Authors: Hernan Alonso, Peter L. Cummins, Jill E. GreadyAbstract:Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) catalyzes the transfer of the N5-methyl group from N5-methyltetrahydrofolate (CH(3)THF) to the cobalt center of a corrinoid/iron-sulfur protein, a reaction similar to that of cobalamin-dependent methionine synthase (MetH). For such a reaction to occur, CH(3)THF is expected to be activated by a stereospecific Protonation at the N5 position. It has been shown experimentally that binding to MeTr is associated with a pK(a) increase and proton uptake. The enzyme could achieve this by binding the unprotonated form of CH(3)THF, followed by specific Protonation at the correct orientation. Here we have used computational approaches to investigate the Protonation State of the ligand and active-site residues in MeTr. First, quantum mechanical (QM) methods with the PCM solvation model were used to predict Protonation positions and pK(a) values of pterin, folate, and their analogues in an aqueous environment. After a reliable calibration of computational and experimental results was obtained, the effect of the protein environment was then considered. Different Protonation States of CH(3)THF and active-site aspartic residues (D75 and D160) were investigated using QM calculations of active-site fragment complexes and the perturbed quantum atom (PQA) approach within QM/MM simulations. The final free energy results indicate that the N5 position of the tetrahydropterin ring is the preferred Protonation position of CH(3)THF when bound to the active site of MeTr, followed by Asp160. We also found that the active-site environment is likely to increase the pK(a) of N5 by about 3 units, leading to proton uptake upon CH(3)THF binding, as observed experimentally for MeTr. Some implications of the results are discussed for the MetH mechanism.
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methyltetrahydrofolate corrinoid iron sulfur protein methyltransferase metr Protonation State of the ligand and active site residues
Journal of Physical Chemistry B, 2009Co-Authors: Hernan Alonso, Peter L. Cummins, Jill E. GreadyAbstract:Methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase (MeTr) catalyzes the transfer of the N5-methyl group from N5-methyltetrahydrofolate (CH(3)THF) to the cobalt center of a corrinoid/iron-sulfur protein, a reaction similar to that of cobalamin-dependent methionine synthase (MetH). For such a reaction to occur, CH(3)THF is expected to be activated by a stereospecific Protonation at the N5 position. It has been shown experimentally that binding to MeTr is associated with a pK(a) increase and proton uptake. The enzyme could achieve this by binding the unprotonated form of CH(3)THF, followed by specific Protonation at the correct orientation. Here we have used computational approaches to investigate the Protonation State of the ligand and active-site residues in MeTr. First, quantum mechanical (QM) methods with the PCM solvation model were used to predict Protonation positions and pK(a) values of pterin, folate, and their analogues in an aqueous environment. After a reliable calibration of computational and experimental results was obtained, the effect of the protein environment was then considered. Different Protonation States of CH(3)THF and active-site aspartic residues (D75 and D160) were investigated using QM calculations of active-site fragment complexes and the perturbed quantum atom (PQA) approach within QM/MM simulations. The final free energy results indicate that the N5 position of the tetrahydropterin ring is the preferred Protonation position of CH(3)THF when bound to the active site of MeTr, followed by Asp160. We also found that the active-site environment is likely to increase the pK(a) of N5 by about 3 units, leading to proton uptake upon CH(3)THF binding, as observed experimentally for MeTr. Some implications of the results are discussed for the MetH mechanism.
David J D Wilson - One of the best experts on this subject based on the ideXlab platform.
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effect of atomic charge solvation entropy and ligand Protonation State on mm pb gb sa binding energies of hiv protease
Journal of Computational Chemistry, 2012Co-Authors: Daniel P Oehme, Robert T C Brownlee, David J D WilsonAbstract:The molecular mechanics-Poisson-Boltzmann surface area (MM-PBSA) and MM-generalized-Born surface area (MM-GBSA) approaches are commonly used in molecular modeling and drug design. Four critical aspects of these approaches have been investigated for their effect on calculated binding energies: (1) the atomic partial charge method used to parameterize the ligand force field, (2) the method used to calculate the solvation free energy, (3) inclusion of entropy estimates, and (4) the Protonation State of the ligand. HIV protease has been used as a test case with six structurally different inhibitors covering a broad range of binding strength to assess the effect of these four parameters. Atomic charge methods are demonstrated to effect both the molecular dynamics (MD) simulation and MM-PB(GB)SA binding energy calculation, with a greater effect on the MD simulation. Coefficients of determination and Spearman rank coefficients were used to quantify the performance of the MM-PB(GB)SA methods relative to the experimental data. In general, better performance was achieved using (i) atomic charge models that produced smaller mean absolute atomic charges (Gasteiger, HF/STO-3G and B3LYP/cc-pVTZ), (ii) the MM-GBSA approach over MM-PBSA, while (iii) inclusion of entropy had a slightly positive effect on correlations with experiment. Accurate representation of the ligand Protonation State was found to be important. It is demonstrated that these approaches can distinguish ligands according to binding strength, underlining the usefulness of these approaches in computer-aided drug design. © 2012 Wiley Periodicals, Inc.
Shahriar Mobashery - One of the best experts on this subject based on the ideXlab platform.
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the importance of a critical Protonation State and the fate of the catalytic steps in class a β lactamases and penicillin binding proteins
Journal of Biological Chemistry, 2004Co-Authors: Dasantila Golemikotra, Samy O Meroueh, Sergei B Vakulenko, Alexey Bulychev, Ann J Stemmler, Timothy L Stemmler, Shahriar MobasheryAbstract:Abstract β-Lactamases and penicillin-binding proteins are bacterial enzymes involved in antibiotic resistance to β-lactam antibiotics and biosynthetic assembly of cell wall, respectively. Members of these large families of enzymes all experience acylation by their respective substrates at an active site serine as the first step in their catalytic activities. A Ser-X-X-Lys sequence motif is seen in all these proteins, and crystal structures demonstrate that the side-chain functions of the serine and lysine are in contact with one another. Three independent methods were used in this report to address the question of the Protonation State of this important lysine (Lys-73) in the TEM-1 β-lactamase from Escherichia coli. These techniques included perturbation of the pKa of Lys-73 by the study of the γ-thialysine-73 variant and the attendant kinetic analyses, investigation of the Protonation State by titration of specifically labeled proteins by nuclear magnetic resonance, and by computational treatment using the thermodynamic integration method. All three methods indicated that the pKa of Lys-73 of this enzyme is attenuated to 8.0–8.5. It is argued herein that the unique ground-State ion pair of Glu-166 and Lys-73 of class A β-lactamases has actually raised the pKa of the active site lysine to 8.0–8.5 from that of the parental penicillin-binding protein. Whereas we cannot rule out that Glu-166 might activate the active site water, which in turn promotes Ser-70 for the acylation event, such as proposed earlier, we would like to propose as a plausible alternative for the acylation step the possibility that the ion pair would reconfigure to the protonated Glu-166 and unprotonated Lys-73. As such, unprotonated Lys-73 could promote serine for acylation, a process that should be shared among all active-site serine β-lactamases and penicillin-binding proteins.
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the importance of a critical Protonation State and the fate of the catalytic steps in class a β lactamases and penicillin binding proteins
Journal of Biological Chemistry, 2004Co-Authors: Dasantila Golemikotra, Samy O Meroueh, Sergei B Vakulenko, Alexey Bulychev, Ann J Stemmler, Timothy L Stemmler, Choonkeun Kim, Shahriar MobasheryAbstract:Beta-lactamases and penicillin-binding proteins are bacterial enzymes involved in antibiotic resistance to beta-lactam antibiotics and biosynthetic assembly of cell wall, respectively. Members of these large families of enzymes all experience acylation by their respective substrates at an active site serine as the first step in their catalytic activities. A Ser-X-X-Lys sequence motif is seen in all these proteins, and crystal structures demonstrate that the side-chain functions of the serine and lysine are in contact with one another. Three independent methods were used in this report to address the question of the Protonation State of this important lysine (Lys-73) in the TEM-1 beta-lactamase from Escherichia coli. These techniques included perturbation of the pK(a) of Lys-73 by the study of the gamma-thialysine-73 variant and the attendant kinetic analyses, investigation of the Protonation State by titration of specifically labeled proteins by nuclear magnetic resonance, and by computational treatment using the thermodynamic integration method. All three methods indicated that the pK(a) of Lys-73 of this enzyme is attenuated to 8.0-8.5. It is argued herein that the unique ground-State ion pair of Glu-166 and Lys-73 of class A beta-lactamases has actually raised the pK(a) of the active site lysine to 8.0-8.5 from that of the parental penicillin-binding protein. Whereas we cannot rule out that Glu-166 might activate the active site water, which in turn promotes Ser-70 for the acylation event, such as proposed earlier, we would like to propose as a plausible alternative for the acylation step the possibility that the ion pair would reconfigure to the protonated Glu-166 and unprotonated Lys-73. As such, unprotonated Lys-73 could promote serine for acylation, a process that should be shared among all active-site serine beta-lactamases and penicillin-binding proteins.
