Akademik Veri Yönetim Sistemi
Uluslararası Biyoteknoloji Kongresi, İstanbul, Türkiye, 28 - 30 Eylül 2023, ss.75-76
Introduction: Glucansucrases are members of the glycoside hydrolase 70 (GH70) enzyme family, which synthesizes polysaccharides and oligosaccharides using sucrose as a substrate. These enzymes are generally responsible for the formation of different glucans, and this difference is due to the α-(1→6) bonds in the main chain and the varying amounts of α-(1→2), α-(1→3) and α-(1→4) bonds in the chain. Weissella confusa can be isolated from different environments such as humans, animals, fermented foods and beverages, grains and vegetables. In previous studies, it has been stated that Weissella confusa strains have high glucansucrase activity and the enzymes responsible for production were determined. Even if Weissella strains have similar glucansucrases, different polysaccharides and oligosaccharides produced may have different functional and technological properties. In this study a novel glucansucrase from sourdough isolate Weissella confusa SDE was identified, heterologously expressed and the structure of glucan SDE was determined.
Materials and methods: Weissella confusa SDE was isolated and cultivated. Whole genome sequencing of Weissella confusa SDE was performed and the similarities between glucansucrase SDE and other glucansucrases available in the NCBI database were identified using NCBI BLASTp searches. Multiple amino acid sequence alignments were conducted with Clustal Omega Tool. The aLICator Ligation Independent Cloning and Expression Kit (Thermo Scientific, USA) was used to clone the glucansucrase gene from Weissella confusa SDE. This gene was cloned into the IPTG-inducible vector pLATE31 and expressed in the host E. coli BL21 (DE3). For the production of glucan with glucansucrase SDE enzyme, the reaction medium was established with sucrose and the resulting glucan was lyophilized. Structural, molecular and thermal characterization of this glucan conducted with the 1H NMR analysis, Fourier transform infrared (FTIR) spectroscopy, differential scanning calorimeter (DSC) analysis, respectively.
Results: Under in vitro conditions, a glucan SDE was produced by glucansucrase SDE using sucrose as the substrate. FTIR analysis of glucan SDE revealed the presence of functional groups within the structure, suggesting that this polymer is a polysaccharide. According to the results of DSC analysis, the degradation of glucan SDE started around 200°C and the melting point of glucan was determined to be approximately 233 °C. Overall, 1H NMR analysis results showed the production of glucans with α-(1→3) and α-(1→6) glycosidic linkages via the reaction of glucansucrase SDE.
Conclusions: Weissella spp., which are very widely found in nature and has been limited studied so far, it is important to isolate its new glucansucrases, expression of recombinant enzymes, determination of their characteristic properties and investigation of their potentials. In this study, these findings showed the production of glucan with α-(1→3) and α-(1→6) glycosidic linkages. The FTIR analysis showed the presence of the functional groups within the glucan structure. The thermal properties of glucan SDE may enable its use in many food matrices, including bakery products. Consequently, the physicochemical properties of glucan SDE would make it a potential candidate for applications in the food industry.
Keywords: Glucansucrase, glucan, Weissella