Ab Initio QM/MM Free-Energy Studies of Arginine Deiminase Catalysis: The Protonation State of the Cys Nucleophile

Ke, Zhihong; Guo, Hua; Xie, Daiqian; Wang, Shenglong; Zhang, Yingkai

doi:10.1021/jp200843s

Cited by 20 publications

(21 citation statements)

References 64 publications

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“…Finally, 10 ns MD simulation for equilibration was performed. [51][52][53][54][55][56][57][58][59][60] As for the MM subsystem, the Amber99SB force field [35][36][37] was employed as in the above classical MD simulations. The Langevin method was utilized to control the temperature at 300 K. The bonds involving hydrogen atoms were constrained by using the SHAKE scheme.…”

Section: Methodsmentioning

confidence: 99%

A full picture of enzymatic catalysis by hydroxynitrile lyases from Hevea brasiliensis: protonation dependent reaction steps and residue-gated movement of the substrate and the product

Zhao

Chen

et al. 2014

Phys. Chem. Chem. Phys.

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Hydroxynitrile lyases (HNLs) defend plants from herbivores and microbial attack by releasing cyanide from hydroxynitriles. The reverse process has been productively applied to bioorganic syntheses of pharmaceuticals and agrochemicals. To improve our understanding of the catalytic mechanism of HNLs, extensive ab initio QM/MM and classical MM molecular dynamics simulations have been performed to explore the catalytic conversion of cyanohydrins into aldehyde (or ketone) and HCN by hydroxynitrile lyases from Hevea brasiliensis (HbHNLs). It was found that the catalytic reaction approximately follows a two-stage mechanism. The first stage involves two fast processes including the proton abstraction of the substrate through a double-proton transfer and the C-CN bond cleavage, while the second stage concerns HCN formation and is rate-determining. The complete free energy profile exhibits a peak of ∼18 kcal mol(-1). Interestingly, the protonation state of Lys236 influences the efficiency of the enzyme only to some extent, but it changes the entire catalytic mechanism. The dynamical behaviors of substrate delivery and HCN release are basically modulated by the gate movement of Trp128. The remarkable exothermicity of substrate binding and the facile release of HCN may drive the enzyme-catalyzed reaction to proceed along the substrate decomposition efficiently. Computational mutagenesis reveals the key residues which play an important role in the catalytic process.

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Section: Methodsmentioning

confidence: 99%

A full picture of enzymatic catalysis by hydroxynitrile lyases from Hevea brasiliensis: protonation dependent reaction steps and residue-gated movement of the substrate and the product

Zhao

Chen

et al. 2014

Phys. Chem. Chem. Phys.

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“…further explored the inhibition mechanism of PAD4 by F‐amidine and the protonation state of active site cysteine . They found that the cysteine and histidine exist as a thiolate–imidazolium ion pair, which was inconsistent with the results of Ke et al . In addition, study of ADI enzyme supports the ionic state of active site .…”

Section: Introductionmentioning

confidence: 87%

“…For example, in the proposed deamination mechanism, the three enzymes (PPAD, PAD4 and ADI) employ the same mechanism. However, the active site of PPAD is clearly different from PAD4 and ADI . And during the hydrolysis stage of PPAD, D238 residue was supposed to only play a role in enhancing the nucleophilic ability of hydrolytic water .…”

Section: Introductionmentioning

confidence: 99%

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Theoretical insights into the protonation states of active site cysteine and citrullination mechanism of Porphyromonas gingivalis peptidylarginine deiminase

et al. 2017

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Porphyromonas gingivalis peptidylarginine deiminase (PPAD) catalyzes the citrullination of peptidylarginine, which plays a critical role in the rheumatoid arthritis (RA) and gene regulation. For a better understanding of citrullination mechanism of PPAD, it is required to establish the protonation states of active site cysteine, which is still a controversial issue for the members of guanidino-group-modifying enzyme superfamily. In this work, we first explored the transformation between the two states: State N (both C351 and H236 are neutral) and State I (both residues exist as a thiolate-imidazolium ion pair), and then investigated the citrullination reaction of peptidylarginine, using a combined QM/MM approach. State N is calculated to be more stable than State I by 8.46 kcal/mol, and State N can transform to State I via two steps of substrate-assisted proton transfer. Citrullination of the peptidylarginine contains deamination and hydrolysis. Starting from State N, the deamination reaction corresponds to an energy barrier of 18.82 kcal/mol. The deprotonated C351 initiates the nucleophilic attack to the substrate, which is the key step for deamination reaction. The hydrolysis reaction contains two chemical steps. Both the deprotonated D238 and H236 can act as the bases to activate the hydrolytic water, which correspond to similar energy barriers (∼17 kcal/mol). On the basis of our calculations, C351, D238, and H236 constitute a catalytic triad, and their protonation states are critical for both the deamination and hydrolysis processes. In view of the sequence similarity, these findings may be shared with human PAD1-PAD4 and other guanidino-group-modifying enzymes. Proteins 2017; 85:1518-1528. © 2017 Wiley Periodicals, Inc.

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“…The Berendsen thermostat 40 was used to control the system temperature at 300 K. The final PMF was calculated with the weighted histogram analysis method (WHAM). 41 The QM/MM method used here is essentially the same as in our recent publication on arginine deiminase, 42 which is a member of the same enzyme superfamily as DDAH and is believed to have a similar catalytic mechanism. 43,44 …”

Section: Methodsmentioning

confidence: 99%

On the Mechanism of Dimethylarginine Dimethylaminohydrolase Inactivation by 4-Halopyridines

Johnson

Monzingo²,

Ke³

et al. 2011

J. Am. Chem. Soc.

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Small molecules capable of selective covalent protein modification are of significant interest for the development of biological probes and therapeutics. We recently reported that 2-methyl-4-bromopyridine is a quiescent affinity label for the nitric oxide controlling enzyme dimethylarginine dimethylaminohydrolase (DDAH) Am. Chem. Soc. 133, 1553-1562. Discovery of this novel protein modifier raised the possibility that the 4-halopyridine motif may be suitable for wider application. Therefore, the inactivation mechanism of the related compound 2-hydroxymethyl-4-chloropyridine is probed here in more detail. Solution studies support an inactivation mechanism in which the active-site Asp66 residue stabilizes the pyridinium form of the inactivator, which has enhanced reactivity toward the active site Cys, resulting in covalent bond formation, loss of the halide, and irreversible inactivation. A 2.18 Å resolution X-ray crystal structure of the inactivated complex elucidates the orientation of the inactivator and its covalent attachment to the active-site Cys, but the structural model does not show an interaction between the inactivator and Asp66. Molecular modeling is used to investigate inactivator binding, reaction, and also a final pyridinium deprotonation step that accounts for the apparent differences between the solution-based and structural studies with respect to the role of Asp66. This work integrates multiple approaches to elucidate the inactivation mechanism of a novel 4-halopyridine "warhead," emphasizing the strategy of using pyridinium formation as a "switch" to enhance reactivity when bound to the target protein.[Small molecules that are capable of covalently modifying proteins are currently undergoing a type of renaissance as they are newly applied to solve problems in chemical biology and proteomics, as well as in drug design, discovery and development. [1][2][3] There are a number of strategies that incorporate moderately electrophilic groups into the design of affinity-based probes, activity-based probes and activity-based protein profiling reagents. A few noted examples include the use of electrophilic phosphonates to target serine hydrolases, 4 * To whom correspondence should be addressed. W.F.: College of Pharmacy, PHAR-MED CHEM, 1 University Station; C0850, Austin, Texas 78712; Phone: (512) Fax: (512) During an effort to find novel inhibitors of the nitric oxide-controlling enzyme dimethylarginine dimethylaminohydrolase (DDAH), 11 2-methyl-4-bromopyridine was discovered to be a time-dependent inhibitor. 12 Mechanistic analysis determined that this 4-halopyridine is a covalent inactivator that selectively modifies the active-site Cys residue. The inactivation mechanism is most consistent with that described for quiescent affinity labels 13,14 in that the compound is relatively unreactive (quiescent) to most biological nucleophiles, but demonstrates an enhanced reactivity when bound to the active site of the target enzyme. In the specific case of 2-methyl-4-bromopyridine, DDAH was pro...

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Ab Initio QM/MM Free-Energy Studies of Arginine Deiminase Catalysis: The Protonation State of the Cys Nucleophile

Cited by 20 publications

References 64 publications

A full picture of enzymatic catalysis by hydroxynitrile lyases from Hevea brasiliensis: protonation dependent reaction steps and residue-gated movement of the substrate and the product

A full picture of enzymatic catalysis by hydroxynitrile lyases from Hevea brasiliensis: protonation dependent reaction steps and residue-gated movement of the substrate and the product

Theoretical insights into the protonation states of active site cysteine and citrullination mechanism of Porphyromonas gingivalis peptidylarginine deiminase

On the Mechanism of Dimethylarginine Dimethylaminohydrolase Inactivation by 4-Halopyridines

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