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Theoretical study of the RNA hydrolysis mechanism of the dinuclear zinc enzyme RNase Z
2009 (English)In: European Journal of Inorganic Chemistry, ISSN 1434-1948, E-ISSN 1099-0682, no 20, 2967-2972 p.Article in journal (Refereed) Published
Abstract [en]

 RNase Z is a dinuclear zinc enzyme that catalyzes the removal of the tRNA 3'-end trailer. Density functional theory is used to investigate the phosphodiester hydrolysis mechanism of this enzyme with a model of the active site constructed on the basis of the crystal structure. The calculations imply that the reaction proceeds through two steps. The first step is a nucleophihc attack by a bridging hydroxide coupled with protonation of the leaving group by a Glu-His diad. Subsequently, a water molecule activated by the same Glu-His diad makes a reverse attack, regenerating the bridging hydroxide. The second step is calculated to be the rate-limiting step with a barrier of 18 kcal/mol, in good agreement with experimental kinetic studies. Both zinc ions participate in substrate binding and orientation, facilitating nucleophilic attack. In addition, they act as electrophilic catalysts to stabilize the pentacoordinate trigonal-bipyramidal transition states.

Place, publisher, year, edition, pages
2009. no 20, 2967-2972 p.
Keyword [en]
Enzyme catalysis, Metalloenzymes, Dinuclear zinc enzymes, Density functional calculations, Hydrolysis, Reaction mechanisms, Zinc
National Category
Chemical Sciences
URN: urn:nbn:se:kth:diva-10174DOI: 10.1002/ejic.200900202ISI: 000268290000010OAI: diva2:209853
QC 20101018. Uppdaterad från Submitted till Published (20101018).Available from: 2009-03-27 Created: 2009-03-27 Last updated: 2010-10-18Bibliographically approved
In thesis
1. Mechanistic insights into dinuclear zinc enzymes from density functional theory studies
Open this publication in new window or tab >>Mechanistic insights into dinuclear zinc enzymes from density functional theory studies
2009 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

In this thesis, quantum chemical methods have been used to shed light on the reaction mechanisms of several dinuclear zinc enzymes. The enzymes studied are involved in the hydrolysis of phosphates, amides, and carboxylic esters, namely RNase Z, Dihydroorotase (DHO), and N-acyl homoserine lactone hydrolase (AHL lactonase). The density functional method B3LYP, together with quite large active site models, was used to investigate these enzymatic reactions. Several plausible proposed mechanisms, involving protonation states of important active site residues (DHO), substrate orientations (AHL lactonase), have been considered. The calculated energetics can be used to assess the feasibility of the suggested reaction mechanisms. Based on the calculations and also on other related dinuclear zinc enzymes studied previously, some general mechanistic features have been uncovered.

For all three enzymes, the nucleophilicity of the bridging hydroxide is shown to be sufficient to perform the nucleophilic attack on the substrates. During the attack, the negative charge is transferred from the bridging hydroxide to the substrate oxygen (P=O or C=O). For phosphate hydrolysis, an in line attack have been suggested for RNase Z. In addition, the two zinc ions in RNase Z are directly involved in stabilizing the negative charge in the penta-coordinated transition states. For carbonyl substrates, only one zinc ion participates in the oxygen anion stabilization in the transition states and the tetrahedral intermediates. Furthermore, the enzymes always use the zinc ion with less negatively-charged ligands to play such role.

All the substrates investigated have poor leaving groups. Therefore, either the zinc ions or some active site residues help the cleavage of the scissile bond. For RNase Z, a Glu-His diad was suggested to protonate the leaving group. For DHO, an Asp residue was shown to transfer a proton from the bridging hydroxide to the leaving group nitrogen. For AHL lactonase, a zinc ion was also observed to stabilize the leaving oxygen anion.

Place, publisher, year, edition, pages
Stockholm: KTH, 2009. 59 p.
Trita-BIO-Report, ISSN 1654-2312 ; 2009:4
National Category
Other Basic Medicine
urn:nbn:se:kth:diva-10175 (URN)978-91-7415-261-6 (ISBN)
Available from: 2009-03-27 Created: 2009-03-27 Last updated: 2010-10-18Bibliographically approved

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Liao, Rong-Zhen
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