Molecular Modelling of a Thermostable Glycoside Hydrolase from &lt;i&gt;Caldivirga &lt;/i&gt;&lt;i&gt;m&lt;/i&gt;&lt;i&gt;aquilingensis&lt;/i&gt; and Its Substrate Docking Mechanism for Galactooligosaccharides Synthesis

Rebaone Letsididi,Bo Jiang,Tao Zhang,Wanmeng Mu,Kekgabile S Letsididi

doi:10.12691/bb-6-1-1

Abstract

Glycoside hydrolases (GHs) are very important enzymes that can catalyze the breakdown of the glycosidic bonds between carbohydrates and non-carbohydrates to synthesize GOS prebiotic sugars and hydrolyze lactose in the dairy industry. GOS can stimulate the growth of gut microbiota to have beneficial health effects in the host. The current work investigated molecular modelling and lactose substrate docking of a thermostable GH family 1 from Caldivirga maquilingensis strain IC-167 and determined its mechanism for GOS synthesis. The 3D model structure obtained from the Swiss model analysis tool revealed that GH 1 from Caldivirga maquilingensis was a tetramer with a catalytic pocket at the center of each monomer. The overall quality of the model was 93%. When lactose was docked in the catalytic site using AutoDock Vina software, the catalytic amino acid residues were identified to be Glu 387 and Glu 209 which acted as nucleophile and acid/base residues respectively. Other amino acid residues like His 153, Gln 19, Glu 432, Ala 266, Asn 267, Ser 268 and Trp 433 were also found to be surrounding the catalytic site and playing an essential role in ligand binding and recognition of the lactose substrate for GOS synthesis. These findings offer an understanding of how enzyme protein structure determines catalytic specificity, which serves as new knowledge basis to engineer GH 1 from C. maquilingensis for the biosynthesis of GOS with a broad or narrow degree of polymerization range.

Highlights

Production of non-digestible galacto-oligosaccharides (GOS) has continued to gain importance because of their recognition as prebiotic compounds that stimulate growth of bifidobacteria and lactobacilli bacteria associated with beneficial health effects and as an interesting functional food ingredient [1,2]
It is largely known that the substrate specificity and the mode of action of Glycoside hydrolases (GHs) are driven by details of their three dimensional structures rather than by their global fold [17]
When the GH 1 from C. maquilingensis amino acid sequence was submitted to the swiss model server to search for a template protein structure with the highest similarity to the target, the most similar template structure was a β-glucosidase from Sulfolobus Solfataricus (PDB: 5ixe.4 ), with 74.69% identity as shown on Figure 1

Summary

Introduction

Production of non-digestible galacto-oligosaccharides (GOS) has continued to gain importance because of their recognition as prebiotic compounds that stimulate growth of bifidobacteria and lactobacilli bacteria associated with beneficial health effects and as an interesting functional food ingredient [1,2]. GOS are of special interest because they have structurally related oligosaccharides present in human breast milk [3,4]. Enzymatic synthesis of GOS from lactose is normally conducted using glycoside hydrolases (GH, EC 3.2.1.X) because these enzymes can transfer glycosyl moieties from a donor sugar to an acceptor [5,6]. This method is the one currently used in the dairy industry for GOS synthesis from lactose with the yield of the reaction depending on the relative ratio of the transgalactosylation versus hydrolysis reaction [7,8]. Enzymatic hydrolysis of the glycosidic bond takes place via general acid catalysis that requires two critical residues: a proton donor and a nucleophile/base [9,10]

Methods

Results

Conclusion