A repertoire of alginate lyases in the alginate polysaccharide utilization loci of marine bacterium Wenyingzhuangia fucanilytica: biochemical properties and action pattern

Abstract

BACKGROUNDAlginate lyases are important tools for alginate biodegradation and oligosaccharide production, which have great potential in food and biofuel fields. The alginate polysaccharide utilization loci (PUL) typically encode a series of alginate lyases with the synergistic action pattern. Exploring valuable alginate lyases and revealing the synergistic effect of enzymes in the PUL is of great significance.RESULTSAn alginate PUL was discovered from the marine bacterium Wenyingzhuangia fucanilytica CZ1127T, and a repertoire of alginate lyases within it was cloned, expressed and characterized. The four alginate lyases in PUL demonstrated similar optimal reaction conditions: maximum enzyme activity at 35‐50 °C, pH 8.0‐9.0. The results of action pattern indicated that they were two PL7 endolytic bifunctional enzymes (Aly7A and Aly7B), a PL6 exolytic bifunctional enzyme (Aly6A) and a PL17 exolytic M‐specific enzyme (Aly17A). The UPLC‐MS technique was employed to reveal the synergistic effect of the four enzymes. The end products of Aly7A were further degraded by Aly7B and eventually generated oligosaccharides with disaccharide to heptasaccharide. The oligosaccharide products were completely degraded to monosaccharides by Aly6A, while it was unable to directly degrade alginate. Aly17A could also produce monosaccharides by cleaving the M‐blocks of oligosaccharide products, as well as the M‐blocks of polysaccharides. The combination of these enzymes resulted in the complete degradation of alginate to monosaccharides.CONCLUSIONA new alginate PUL was mined and four novel alginate lyases in the PUL were expressed and characterized. The four cooperative alginate lyases provide novel tools for alginate degradation and biological fermentation.This article is protected by copyright. All rights reserved.

Action Pattern of a Novel G-Specific Alginate Lyase: Determination of Subsite Specificity by HPAEC–PAD/MS

“…The purity and M w of the purified target protein were subsequently assessed by SDS-PAGE. The detailed procedures for the expression and purification were consistent with our previous study …”