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Reaction mechanism of retaining glycosyltransferases - a computational study

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TRNKA Tomáš KOZMON Stanislav TVAROŠKA Igor KOČA Jaroslav

Rok publikování 2013
Druh Další prezentace na konferencích
Fakulta / Pracoviště MU

Středoevropský technologický institut

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Popis Glycosylation of cell surface proteins plays a crucial role in cell communication and recognition. Alterations in glycan structures are linked to many diseases with the most prominent example being cancer. To understand the regulation of glycosylation and to be able to modify it, reaction mechanisms of involved glycosyltransferases need to be known. However, reaction mechanism of the configuration-retaining group of glycosyltransferases hasn't been sufficiently explained yet. For this reason we have chosen a retaining glycosyltransferase – polypeptide UDP-GalNAc transferase (ppGalNAcT) – as the subject of our quantum-chemical study. This enzyme catalyses the transfer of N-acetylgalactosamine moiety onto serine or threonine hydroxyls, forming the first bond of the so-called O-linked glycosylation pathway. Increased activity of ppGalNAcT has been found to enable metastasis of breast and colorectal cancer. We're studying human ppGalNAcT2 by a hybrid QM/MM approach using density functional theory for the important part of the active site. Two-dimensional potential energy surface scan on the OPBE-D3/TZP level was used to obtain initial approximation of the reaction path and location of transition state candidates. Distance-difference coordinates were implemented into the used QM/MM software to overcome the inability of two simple distance coordinates to describe all the relevant degrees of freedom. After obtaining the 2D energy map, further optimisation was carried out in order to refine the minimum energy path and transition states. According to our results, the reaction proceeds via a two-step same-face SNi-like nucleophilic displacement with a very short-lived carbocationic intermediate. The proposed mechanism agrees well with recent experimental evidence. Additionally, to facilitate further studies of the reactivity of this system and related glycosyltransferases, a ReaxFF reactive force field for classical molecular dynamics of biomolecules is being parameterised.
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