Dextran is an α-(1→6)-glucan that is synthesized by some lactic acid bacteria, and branched dextran with α-(1→2)-, α-(1→3)-, and α-(1→4)-linkages are often produced. Although many dextranases are known to act on the α-(1→6)-linkage of dextran, few studies have functionally analyzed the proteins involved in degrading branched dextran. The mechanism by which bacteria utilize branched dextran is unknown. Earlier, we identified dextranase (FjDex31A) and kojibiose hydrolase (FjGH65A) in the dextran utilization locus (FjDexUL) of a soil Bacteroidota Flavobacterium johnsoniae and hypothesized that FjDexUL is involved in the degradation of α-(1→2)-branched dextran. In this study, we demonstrate that FjDexUL proteins recognize and degrade α-(1→2)- and α-(1→3)-branched dextrans produced by Leuconostoc citreum S-32 (S-32 α-glucan). The FjDexUL genes were significantly upregulated when S-32 α-glucan was the carbon source compared with α-glucooligosaccharides and α-glucans, such as linear dextran and branched α-glucan from L. citreum S-64. FjDexUL glycoside hydrolases synergistically degraded S-32 α-glucan. The crystal structure of FjGH66 shows that some sugar-binding subsites can accommodate α-(1→2)- and α-(1→3)-branches. The structure of FjGH65A in complex with isomaltose supports that FjGH65A acts on α-(1→2)-glucosyl isomaltooligosaccharides. Furthermore, two cell surface sugar-binding proteins (FjDusD and FjDusE) were characterized, and FjDusD showed an affinity for isomaltooligosaccharides and FjDusE for dextran, including linear and branched dextrans. Collectively, FjDexUL proteins are suggested to be involved in the degradation of α-(1→2)- and α-(1→3)-branched dextrans. Our results will be helpful in understanding the bacterial nutrient requirements and symbiotic relationships between bacteria at the molecular level.

右旋糖酐是由一些乳酸菌合成的α-(1→6)-葡聚糖，支链右旋糖酐有α-(1→2)-、α-(1→3)-和α-(1→4) )-联系经常产生。尽管已知许多葡聚糖酶作用于葡聚糖的 α-(1→6)-连接，但很少有研究对参与降解支链葡聚糖的蛋白质进行功能分析。细菌利用支链葡聚糖的机制尚不清楚。早些时候，我们在土壤约氏黄杆菌的葡聚糖利用位点（FjDexUL）中鉴定了葡聚糖酶（FjDex31A）和曲二糖水解酶（FjGH65A），并假设FjDexUL参与α-（1→2）-支链葡聚糖的降解。在本研究中，我们证明 FjDexUL 蛋白可识别并降解柠檬明串珠菌S-32（S-32 α-葡聚糖）产生的 α-(1→2)- 和 α-(1→3)- 支链葡聚糖。与 α-低聚葡萄糖和 α-葡聚糖（例如来自柠檬酸乳杆菌S-64 的线性葡聚糖和支链 α-葡聚糖）相比，当 S-32 α-葡聚糖为碳源时，FjDexUL 基因显着上调。FjDexUL 糖苷水解酶可协同降解 S-32 α-葡聚糖。FjGH66的晶体结构表明一些糖结合亚位点可以容纳α-(1→2)-和α-(1→3)-分支。FjGH65A 与异麦芽糖复合物的结构支持 FjGH65A 作用于 α-(1→2)-葡糖基低聚异麦芽糖。此外，还对两种细胞表面糖结合蛋白（FjDusD 和 FjDusE）进行了表征，FjDusD 显示出对低聚异麦芽糖的亲和力，FjDusE 显示出对葡聚糖（包括线性和支化葡聚糖）的亲和力。总的来说，FjDexUL 蛋白被认为参与 α-(1→2)- 和 α-(1→3)- 支链葡聚糖的降解。我们的结果将有助于在分子水平上了解细菌的营养需求和细菌之间的共生关系。