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Reaction Mechanism of the Trinuclear Zinc Enzyme Phospholipase C: A Density Functional Theory Study
Stockholm University, Faculty of Science, Department of Organic Chemistry.
Stockholm University, Faculty of Science, Department of Organic Chemistry.
2010 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 114, no 7, 2533-2540 p.Article in journal (Refereed) Published
Abstract [en]

Phosphatidylcholine-preferring phospholipase C is a trinuclear zinc-dependent phosphodiesterase, catalyzing the hydrolysis of choline phospholipids. In the present study, density functional theory is used to investigate the reaction mechanism of this enzyme. Two possible mechanistic scenarios were considered with a model of the active site designed on the basis of the high resolution X-ray crystal structure of the native enzyme. The calculations show that a Zn1 and Zn3 bridging hydroxide rather than a Zn1 coordinated water molecule performs the nucleophilic attack on the phosphorus center. Simultaneously, Zn2 activates a water molecule to protonate the leaving group. In the following step, the newly generated Zn2 bound hydroxide makes the reverse attack, resulting in the regeneration of the bridging hydroxide. The first step is calculated to be rate-limiting with a barrier of 17.3 kcal/mol, in good agreement with experimental kinetic studies. The zinc ions are suggested to orient the substrate for nucleophilic attack and provide electrostatic stabilization to the dianionic penta-coordinated trigonal bipyramidal transition states, thereby lowering the barrier.

Place, publisher, year, edition, pages
2010. Vol. 114, no 7, 2533-2540 p.
National Category
Chemical Sciences
URN: urn:nbn:se:su:diva-43033DOI: 10.1021/jp910992fISI: 000274578500026OAI: diva2:353198
Available from: 2010-09-24 Created: 2010-09-24 Last updated: 2011-11-23Bibliographically approved
In thesis
1. Quantum Chemical Cluster Modeling of Enzymatic Reactions
Open this publication in new window or tab >>Quantum Chemical Cluster Modeling of Enzymatic Reactions
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The Quantum chemical cluster approach has been shown to be quite powerful and efficient in the modeling of enzyme active sites and reaction mechanisms. In this thesis, the reaction mechanisms of several enzymes have been investigated using the hybrid density functional B3LYP. The enzymes studied include four dinuclear zinc enzymes, namely dihydroorotase, N-acyl-homoserine lactone hydrolase, RNase Z, and human renal dipeptidase, two trinuclear zinc enzymes, namely phospholipase C and nuclease P1, two tungstoenzymes, namely formaldehyde ferredoxin oxidoreductase and acetylene hydratase, aspartate α-decarboxylase, and mycolic acid cyclopropane synthase. The potential energy profiles for various mechanistic scenarios have been calculated and analyzed. The role of the metal ions as well as important active site residues has been discussed.

  In the cluster approach, the effects of the parts of the enzyme that are not explicitly included in the model are taken into account using implicit solvation methods.

  For all six zinc-dependent enzymes studied, the di-zinc bridging hydroxide has been shown to be capable of performing nucleophilic attack on the substrate. In addition, one, two, or even all three zinc ions participate in the stabilization of the negative charge in the transition states and intermediates, thereby lowering the barriers.

  For the two tungstoenzymes, several different mechanistic scenarios have been considered to identify the energetically most feasible one. For both enzymes, new mechanisms are proposed.

  Finally, the mechanism of mycolic acid cyclopropane synthase has been shown to be a direct methyl transfer to the substrate double bond, followed by proton transfer to the bicarbonate.

  From the studies of these enzymes, we demonstrate that density functional calculations are able to solve mechanistic problems related to enzymatic reactions, and a wealth of new insight can be obtained.

Place, publisher, year, edition, pages
Stockholm: Department of Organic Chemistry, Stockholm University, 2010. 76 p.
cluster approach, density functional theory, B3LYP, enzyme, mechanism, dinuclear, trinuclear, zinc, tungsten, hydrolysis, decarboxylation, formaldehyde oxidation, hydration, methyl transfer
National Category
Theoretical Chemistry
Research subject
Organic Chemistry
urn:nbn:se:su:diva-43026 (URN)978-91-7447-129-8 (ISBN)
Public defence
2010-10-27, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Available from: 2010-10-05 Created: 2010-09-24 Last updated: 2010-10-07Bibliographically approved

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Liao, Rong-ZhenHimo, Fahmi
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