Effect of a single point mutation on the interaction of glucans with a glucansucrase from Leuconostoc mesenteroides NRRL B-1118

Authors: Cote, Gregory; Skory, Christopher - Chris

Submitted to: Carbohydrate Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/8/2016
Publication Date: 4/28/2016
Citation: Cote, G.L., Skory, C.D. 2016. Effect of a single point mutation on the interaction of glucans with a glucansucrase from Leuconostoc mesenteroides NRRL B-1118. Carbohydrate Research. 428:57-61.

Interpretive Summary: Certain bacteria used in fermented food produce an enzyme called glucansucrase that are able to produce long polymers of glucose from cane or beet sugars. These polymers are typically water-soluble, but we identified a novel glucansucrase enzyme that produces a unique gel-like polymer that is insoluble in water. In this study, showed that the enzymes can interact with soluble polymers to form novel hybrids, and studied the mechanism of these interactions. These polymers have potential for production of biodegradable fibers and films that can be used in a broad number of consumer applications and provide the foundation for developing new eco-friendly materials derived from renewable agricultural materials.

Technical Abstract: Our previous work showed that substitution of an amino acid that is coupled with the +2 subsite adjacent to the transition stabilizer of a glucansucrase, which produces a water-insoluble glucan, resulted in significant changes in the structures and yields of the water-insoluble glucans produced. We now describe how these changes affect the ability of the glucansucrase to bind to exogenous glucans, and how these glucans can influence the yield, product structures, and kinetics of the mutant glucansucrases. The activity of the wild-type enzyme, with threonine at position 654, is not significantly activated by added dextran, and the yield of water-insoluble glucan from sucrose is only slightly increased by dextran. Mutant T654Y is not affected at all by the addition of dextran. However, several mutants exhibit markedly lower yields of glucan relative to the wild type; these lower yields can be partially or completely overcome by the addition of water-soluble dextran. Although evidence indicates that the soluble dextran is incorporated into water-insoluble glucan, the increased yields cannot be accounted for solely by incorporation of the dextran into insoluble product. Furthermore, these mutants are significantly activated by exogenous glucans. The addition of dextran does not markedly change the (KM) for sucrose in the mutants, but does increase the Vmax of the reaction. These effects apparently depend on the presence of unbranched sequences of a1-6-linked D-glucose units in the glucan. Mutations at threonine-654 appear not to affect binding to dextran, but may influence the conformation of the enzyme.