DataSheet1_Low Molar Mass Dextran: One-Step Synthesis With Dextransucrase by Site-Directed Mutagenesis and its Potential of Iron-Loading.docx

Iron dextran is a common anti-anemia drug, and it requires low molar mass dextran as substrate. In this work, we selected 11 amino acid residues in domain A/B of DSR-MΔ2 within a 5-angstrom distance from sucrose for site-directed mutagenesis by molecular docking. Mutation of Q634 did not affect the enzyme catalytic activity, but showed an obvious impact on the ratio of low molecular weight dextran (L-dextran, 3,000–5,000 Da) and relatively higher molecular weight dextran (H-dextran, around 10,000 Da). L-dextran was the main product synthesized by DSR-MΔ2 Q634A, and its average molecular weight was 3,951 Da with a polydispersity index <1.3. The structural characterization of this homopolysaccharide revealed that it was a dextran, with 86.0% α(1→6) and 14.0% α(1→4) glycosidic linkages. Moreover, L-dextran was oxidized with NaOH and chelated with ferric trichloride, and an OL-dextran-iron complex was synthesized with a high iron-loading potential of 33.5% (w/w). Altogether, mutation of amino acids near the sucrose binding site of dextransucrase can affect the chain elongation process, making it possible to modulate dextran size.

右旋糖酐铁（Iron dextran）是一类常用的抗贫血药物，其合成需以低摩尔质量右旋糖酐作为底物。本研究通过分子对接，选取蔗糖结合位点5埃范围内、位于DSR-MΔ2的A/B结构域中的11个氨基酸残基进行定点诱变。Q634位点的突变未对酶的催化活性造成影响，但显著改变了低分子量右旋糖酐（L-右旋糖酐，3000~5000 Da）与相对高分子量右旋糖酐（H-右旋糖酐，约10000 Da）的产物比例。由DSR-MΔ2 Q634A突变体合成的产物以L-右旋糖酐为主，其平均分子量为3951 Da，多分散性指数小于1.3。对该同多糖的结构表征显示，其为右旋糖酐，包含86.0%的α(1→6)糖苷键与14.0%的α(1→4)糖苷键。此外，研究人员通过氢氧化钠氧化、三氯化铁螯合处理该L-右旋糖酐，合成了载铁潜力达33.5%（质量比，w/w）的OL-右旋糖酐-铁复合物。综上，葡聚糖蔗糖酶蔗糖结合位点附近的氨基酸突变可影响多糖链的延伸过程，从而实现右旋糖酐分子量的调控。