Myoglobin has important biological functions in storing and transporting small diatomic molecules in human body. Two possible orientations of carbon monoxide （CO） in the heme distal pocket （named as BI and B2 state...Myoglobin has important biological functions in storing and transporting small diatomic molecules in human body. Two possible orientations of carbon monoxide （CO） in the heme distal pocket （named as BI and B2 states） of myoglobin have been experimentally indicated. In this study, ab initio quantum mechanics/molecular mechanics （QM/MM） molecular dynamics simulation of CO in myoglobin was carried out to investigate the two possible B states. Our results demonstrate that the B1 and B2 states correspond to Fe... CO （with carbon atom closer to iron center of heme） and Fe... OC （with oxygen atom closer to Fe）, by comparing with the experimental infrared spectrum. QM electrostatic polarization effect on CO brought from the protein and solvent environment is the main driving force, which anchors CO in two distinctive orientations and hinders its rotation. The calculated vibrational frequency shift between the state B1 and B2 is 13.1 cm-1, which is in good agreement with experimental value of 11.5 cm-1. This study also shows that the electric field produced by the solvent plays an important role in assisting protein functions by exerting directional electric field at the active site of the protein, From residue-based electric field decomposition, several residues were found to have most contributions to the total electric field at the CO center, including a few charged residues and three adjacent uncharged polar residues （namely, HIS64, ILE107, and PHE43）. This study provides new physical insights on rational design of enzyme with higher electric field at the active site.展开更多
Formate oxidase(FOx),which contains 8-formyl flavin adenine dinucleotide(FAD),exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-cho...Formate oxidase(FOx),which contains 8-formyl flavin adenine dinucleotide(FAD),exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-choline(GMC)oxidoreductase enzymes that contain only the standard FAD cofactor.The FOx-mediated conversion of FAD to 8-formyl FAD results in an approximate 10-fold increase in formate oxidase activity.However,the mechanistic details underlying the autocatalytic formation of 8-formyl FAD are still not well understood,which impedes further utilization of FOx.In this study,we employ molecular dynamics simulation,QM/MM umbrella sampling simulation,enzyme activity assay,site-directed mutagenesis,and spectroscopic analysis to elucidate the oxidation mechanism of FAD to 8-formyl FAD.Our results reveal that a catalytic water molecule,rather than any catalytic amino acids,serves as a general base to deprotonate the C8 methyl group on FAD,thus facilitating the formation of a quinone-methide tautomer intermediate.An oxygen molecule subsequently oxidizes this intermediate,resulting in a C8 methyl hydroperoxide anion that is protonated and dissociated to form OHC-RP and OH−.During the oxidation of FAD to 8-formyl FAD,the energy barrier for the rate-limiting step is calculated to be 22.8 kcal/mol,which corresponds to the required 14-hour transformation time observed experimentally.Further,the elucidated oxidation mechanism reveals that the autocatalytic formation of 8-formyl FAD depends on the proximal arginine and serine residues,R87 and S94,respectively.Enzymatic activity assay validates that the mutation of R87 to lysine reduces the kcat value to 75%of the wild-type,while the mutation to histidine results in a complete loss of activity.Similarly,the mutant S94I also leads to the deactivation of enzyme.This dependency arises because the nucleophilic OH−group and the quinone-methide tautomer intermediate are stabilized through the noncovalent interaction provided by R87 and S94.These findings not only explain the mechanistic details of each reaction step but also clarify the functional role of R87 and S94 during the oxidative maturation of 8-formyl FAD,thereby providing crucial theoretical support for the development of novel flavoenzymes with enhanced redox properties.展开更多
Various computational methods were employed to investigate the zwitterion formation,a critical step for the reaction of monoethanolamine with CO2,in five solvents(water,monoethanolamine,propylamine,methanol and chloro...Various computational methods were employed to investigate the zwitterion formation,a critical step for the reaction of monoethanolamine with CO2,in five solvents(water,monoethanolamine,propylamine,methanol and chloroform)to probe the effect of hydrogen bond capacity of solvents on the reaction of amine with CO2 occurring in the amine-based CO2 capture process.The results indicate that the zwitterion can be formed in all considered solvents except chloroform.For two pairs of solvents(methanol and monoethanolamine,propylamine and chloroform)with similar dielectric constant but different hydrogen bond capacity,the solvents with higher hydrogen bond capacity(monoethanolamine and propylamine)facilitate the zwitterion formation.More importantly,kinetics parameters such as activation free energy for the zwitterion formation are more relevant to the hydrogen bond capacity than to dielectric constant of the considered solvents,clarifying the hydrogen bond capacity could be more important than dielectric constant in determining the kinetics of monoethanolamine with CO2.展开更多
Motivated by several long-lasting mechanistic questions for biomolecular proton pumps,we have engaged in developing hybrid quantum mechanical/molecular mechanical(QM/MM) methods that allow an efficient and reliable de...Motivated by several long-lasting mechanistic questions for biomolecular proton pumps,we have engaged in developing hybrid quantum mechanical/molecular mechanical(QM/MM) methods that allow an efficient and reliable description of long-range proton transport in transmembrane proteins.In this review,we briefly discuss several relevant issues:the need to develop a "multi-scale" generalized solvent boundary potential(GSBP) for the analysis of chemical events in large trans-membrane proteins,approaches to validate such a protocol,and the importance of improving the flexibility of QM/MM Hamiltonian.Several recent studies of model and realistic protein systems are also discussed to help put the discussions into context.Collectively,these studies suggest that the QM/MM-GSBP framework based on an approximate density functional theory(SCC-DFTB) as QM holds the promise to strike the proper balance between computational efficiency,accuracy and generality.With additional improvements in the methodology and recent developments by others,especially powerful sampling techniques,this "multi-scale" framework will be able to help unlock the secrets of proton pumps and other biomolecular machines.展开更多
文摘Myoglobin has important biological functions in storing and transporting small diatomic molecules in human body. Two possible orientations of carbon monoxide （CO） in the heme distal pocket （named as BI and B2 states） of myoglobin have been experimentally indicated. In this study, ab initio quantum mechanics/molecular mechanics （QM/MM） molecular dynamics simulation of CO in myoglobin was carried out to investigate the two possible B states. Our results demonstrate that the B1 and B2 states correspond to Fe... CO （with carbon atom closer to iron center of heme） and Fe... OC （with oxygen atom closer to Fe）, by comparing with the experimental infrared spectrum. QM electrostatic polarization effect on CO brought from the protein and solvent environment is the main driving force, which anchors CO in two distinctive orientations and hinders its rotation. The calculated vibrational frequency shift between the state B1 and B2 is 13.1 cm-1, which is in good agreement with experimental value of 11.5 cm-1. This study also shows that the electric field produced by the solvent plays an important role in assisting protein functions by exerting directional electric field at the active site of the protein, From residue-based electric field decomposition, several residues were found to have most contributions to the total electric field at the CO center, including a few charged residues and three adjacent uncharged polar residues （namely, HIS64, ILE107, and PHE43）. This study provides new physical insights on rational design of enzyme with higher electric field at the active site.
基金supported by the National Natural Science Foundation of China(32201030,32271319 and 32071267)the Science and Technology Department of Jilin Province(20230402041GH and YDZJ202301ZYTS537)+2 种基金the Education Department of Jilin Province(JJKH20220970KJ)the Development and Reform Commission of Jilin Province(2023C015)the Fundamental Research Funds of the Central Universities in China(2024-JCXK-11).
文摘Formate oxidase(FOx),which contains 8-formyl flavin adenine dinucleotide(FAD),exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-choline(GMC)oxidoreductase enzymes that contain only the standard FAD cofactor.The FOx-mediated conversion of FAD to 8-formyl FAD results in an approximate 10-fold increase in formate oxidase activity.However,the mechanistic details underlying the autocatalytic formation of 8-formyl FAD are still not well understood,which impedes further utilization of FOx.In this study,we employ molecular dynamics simulation,QM/MM umbrella sampling simulation,enzyme activity assay,site-directed mutagenesis,and spectroscopic analysis to elucidate the oxidation mechanism of FAD to 8-formyl FAD.Our results reveal that a catalytic water molecule,rather than any catalytic amino acids,serves as a general base to deprotonate the C8 methyl group on FAD,thus facilitating the formation of a quinone-methide tautomer intermediate.An oxygen molecule subsequently oxidizes this intermediate,resulting in a C8 methyl hydroperoxide anion that is protonated and dissociated to form OHC-RP and OH−.During the oxidation of FAD to 8-formyl FAD,the energy barrier for the rate-limiting step is calculated to be 22.8 kcal/mol,which corresponds to the required 14-hour transformation time observed experimentally.Further,the elucidated oxidation mechanism reveals that the autocatalytic formation of 8-formyl FAD depends on the proximal arginine and serine residues,R87 and S94,respectively.Enzymatic activity assay validates that the mutation of R87 to lysine reduces the kcat value to 75%of the wild-type,while the mutation to histidine results in a complete loss of activity.Similarly,the mutant S94I also leads to the deactivation of enzyme.This dependency arises because the nucleophilic OH−group and the quinone-methide tautomer intermediate are stabilized through the noncovalent interaction provided by R87 and S94.These findings not only explain the mechanistic details of each reaction step but also clarify the functional role of R87 and S94 during the oxidative maturation of 8-formyl FAD,thereby providing crucial theoretical support for the development of novel flavoenzymes with enhanced redox properties.
基金supported by the National Natural Science Foundation of China(Nos.21876024 and 21677028)the Major International(Regional)Joint Research Project(No.21661142001)+2 种基金the Program for Changjiang Scholars and Innovative Research Team in University(No.IRT_13R05)the Programme of Introducing Talents of Discipline to Universities(No.B13012)Supercomputing Center of Dalian University of Technology
文摘Various computational methods were employed to investigate the zwitterion formation,a critical step for the reaction of monoethanolamine with CO2,in five solvents(water,monoethanolamine,propylamine,methanol and chloroform)to probe the effect of hydrogen bond capacity of solvents on the reaction of amine with CO2 occurring in the amine-based CO2 capture process.The results indicate that the zwitterion can be formed in all considered solvents except chloroform.For two pairs of solvents(methanol and monoethanolamine,propylamine and chloroform)with similar dielectric constant but different hydrogen bond capacity,the solvents with higher hydrogen bond capacity(monoethanolamine and propylamine)facilitate the zwitterion formation.More importantly,kinetics parameters such as activation free energy for the zwitterion formation are more relevant to the hydrogen bond capacity than to dielectric constant of the considered solvents,clarifying the hydrogen bond capacity could be more important than dielectric constant in determining the kinetics of monoethanolamine with CO2.
基金supported in part by NIH grant R01-GM084028NSF grant CHE-0957285+1 种基金U.S.Department of Energy Genomics:GTL and Sci-DAC Programs (DEFG02-04ER25627)supported in part by the National Science Foundation through a major instrumentation grant (CHE-0840494)
文摘Motivated by several long-lasting mechanistic questions for biomolecular proton pumps,we have engaged in developing hybrid quantum mechanical/molecular mechanical(QM/MM) methods that allow an efficient and reliable description of long-range proton transport in transmembrane proteins.In this review,we briefly discuss several relevant issues:the need to develop a "multi-scale" generalized solvent boundary potential(GSBP) for the analysis of chemical events in large trans-membrane proteins,approaches to validate such a protocol,and the importance of improving the flexibility of QM/MM Hamiltonian.Several recent studies of model and realistic protein systems are also discussed to help put the discussions into context.Collectively,these studies suggest that the QM/MM-GSBP framework based on an approximate density functional theory(SCC-DFTB) as QM holds the promise to strike the proper balance between computational efficiency,accuracy and generality.With additional improvements in the methodology and recent developments by others,especially powerful sampling techniques,this "multi-scale" framework will be able to help unlock the secrets of proton pumps and other biomolecular machines.
