Tautomerization in the UDP-Galactopyranose Mutase Mechanism: A DFT-Cluster and QM/MM Investigation
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- 作者：WenJuan Huang ; James W. Gauld
- 刊名：The Journal of Physical Chemistry B
- 出版年：2012
- 出版时间：December 6, 2012
- 年：2012
- 卷：116
- 期：48
- 页码：14040-14050
- 全文大小：630K
- 年卷期：v.116,no.48(December 6, 2012)
- ISSN：1520-5207
文摘
UDP-galactopyranose mutase (UGM) is a key flavoenzyme involved in cell wall biosynthesis of a variety of pathogenic bacteria and hence, integral to their survival. It catalyzes the interconversion of UDP-galactopyranose (UDP-Galp) and UDP-galactofuranose (UDP-Galf); interconversion of the galactose moieties six- and five-membered ring forms. We have synergistically applied both density functional theory (DFT)-cluster and ONIOM quantum mechanics/molecular mechanics (QM/MM) hybrid calculations to elucidate the mechanism of this important enzyme and to provide insight into its uncommon mechanism. It is shown that the flavin must initially be in its fully reduced form. Furthermore, it requires an N5FAD鈥揌 proton, which, through a series of tautomerizations, is transferred onto the ring oxygen of the substrate鈥檚 Galp moiety to facilitate ring-opening with concomitant Schiff base formation. Conversely, Galf formation is achieved via a series of tautomerizations involving proton transfer from the galactose鈥檚 鈭扥4GalH group ultimately onto the flavin鈥檚 N5FAD center. With the DFT-cluster model, the overall rate-limiting step with a barrier of 120.0 kJ mol鈥? is the interconversion of two Galf-flavin tautomers: one containing a C4FAD鈥揙H group and the other a tetrahedral protonated-N5FAD center. In contrast, in the QM/MM model a considerably more extensive chemical model was used that included all of the residues surrounding the active site, and modeled both their steric and electrostatic effects. In this approach, the overall rate-limiting step with a barrier of 99.2 kJ mol鈥? occurs during conformational rearrangement of the Schiff base linear galactose鈥揻lavin complex. This appears due to the lack of suitable functional groups to facilitate the rearrangement.