Computational analysis on protein-ligand interaction of xylitol-phosphate dehydrogenase enzymes for xylitol production
Xylitol is a high-value low-calorie sweetener used as sugar substitute in food and pharmaceutical industry. Xylitol phosphate dehydrogenase (XPDH) catalyses the conversion of D-xylulose 5-phosphate (XU5P) and D-ribulose 5-phosphate (RU5P) to xylitol and ribitol respectively in the presence of nicoti...
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| Main Author: | |
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| Format: | Thesis |
| Language: | English |
| Published: |
2018
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| Subjects: | |
| Online Access: | http://eprints.utm.my/id/eprint/81420/1/SitiAisyahRazaliPFS2018.pdf |
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| Summary: | Xylitol is a high-value low-calorie sweetener used as sugar substitute in food and pharmaceutical industry. Xylitol phosphate dehydrogenase (XPDH) catalyses the conversion of D-xylulose 5-phosphate (XU5P) and D-ribulose 5-phosphate (RU5P) to xylitol and ribitol respectively in the presence of nicotinamide adenine dinucleotide hydride (NADH). Although these enzymes have been shown to produce xylitol, however there is a limited understanding of the mechanism of the catalytic events of these reactions and the detailed mechanism has yet to be elucidated. Understanding of the catalytic activity of these enzymes would provide novel information for protein engineering to improve xylitol production. The main goal of this work is to analyse the conformational changes of XPDH-bound ligands such as Zn2^ NADH, XU5P, and RU5P to elucidate the key amino acids involved in the substrate binding. In silico modelling, comparative molecular dynamic simulations, interaction analysis and conformational study were carried out on three XPDH enzymes of the Medium-chain dehydrogenase (MDR) family; XPDH from Lactobacillus rhamnosus (LrXPDH) and Clostridium difficile (CdXPDH, Cd1XPDH) in order to elucidate the atomistic details of conformational transition, especially on the open and closed state of XPDH. The critical residues involved in substrate binding and conformational changes were mutated using in silico site-directed mutagenesis. The result showed that residues Cys37, His58, Glu59, and Glu142 form an active site pocket within the catalytic domain. In the coenzyme domain, NADH is shown to bind to highly conserved glycine-rich motif; GXGXXG (residues 166-171). The results also revealed that XPDH consists of a dual mechanism that can catalyse hydride transfer to dissimilar substrates (XU5P and RU5P), which His58 and Ser39 would act as the proton donor for reduction of XU5P and RU5P respectively. The structural comparison and MD simulations displayed a significant difference in the conformational dynamics of the catalytic and coenzyme loops between Apo and XPDH-complexes and highlight the contribution of newly found triad residues (W48, I259, and W285). The study also identified the effect of S39A and W285A mutations on substrate binding and conformational changes. The study successfully elucidated the mechanistic aspect of catalysis mechanism and dynamical event of XPDH enzymes at molecular level. The results from this study would assist future mutagenesis study and enzyme modification work to increase the catalysis efficiency of xylitol production in the industry. |
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