一种新型海洋壳聚糖酶的表达与生化特性研究 适合制备壳二糖。
Expression and Biochemical Characterization of a Novel Marine Chitosanase from Suitable for Preparation of Chitobiose.
机构信息
School of Chemistry and Chemical Engineering, University of Jinan, Jinan 250022, China.
National Key Laboratory of Biochemical Engineering, National Engineering Research Center for Biotechnology (Beijing), Key Laboratory of Biopharmaceutical Production & Formulation Engineering, PLA, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, China.
出版信息
Mar Drugs. 2021 May 24;19(6):300. doi: 10.3390/md19060300.
It is known that bioactivities of chitooligosaccharide (COS) are closely related to the degree of polymerization (DP); therefore, it is essential to prepare COS with controllable DP, such as chitobiose showing high antioxidant and antihyperlipidemia activities. In this study, BLAST, sequence alignment and phylogenetic analysis of characterized glycoside hydrolase (GH) 46 -chitosanases revealed that a chitosanase Sn1-CSN from was different from others. Sn1-CSN was overexpressed in , purified and characterized in detail. It showed the highest activity at pH 6.0 and exhibited superior stability between pH 4.0 and pH 11.0. Sn1-CSN displayed the highest activity at 50 °C and was fairly stable at ≤45 °C. Its apparent kinetic parameters against chitosan (DDA: degree of deacetylation, >94%) were determined, with and values of 1.8 mg/mL and 88.3 s, respectively. Cu enhanced the activity of Sn1-CSN by 54.2%, whereas Fe inhibited activity by 15.1%. Hydrolysis products of chitosan (DDA > 94%) by Sn1-CSN were mainly composed of chitobiose (87.3%), whereas partially acetylated chitosan with DDA 69% was mainly converted into partially acetylated COS with DP 2-13. This -chitosanase has great potential to be used for the preparation of chitobiose and partially acetylated COS with different DPs.
已知壳寡糖（COS）的生物活性与其聚合度（DP）密切相关；因此，制备具有可控 DP 的 COS 至关重要，例如具有高抗氧化和抗高血脂活性的壳二糖。在这项研究中，通过对特征性糖苷水解酶（GH）46-壳聚糖酶的 BLAST、序列比对和系统发育分析，发现来自 的壳聚糖酶 Sn1-CSN 与其他酶不同。Sn1-CSN 在 中过量表达，并进行了详细的纯化和表征。它在 pH6.0 时表现出最高的活性，在 pH4.0 到 pH11.0 之间具有优异的稳定性。Sn1-CSN 在 50°C 时表现出最高的活性，在≤45°C 时相当稳定。确定了其对壳聚糖（DDA：脱乙酰度，>94%）的表观动力学参数， 值和 值分别为 1.8mg/mL 和 88.3s。Cu 增强了 Sn1-CSN 的活性，增加了 54.2%，而 Fe 抑制了 15.1%的活性。Sn1-CSN 水解壳聚糖（DDA>94%）的产物主要由壳二糖（87.3%）组成，而 DDA 为 69%的部分乙酰化壳聚糖主要转化为 DP 为 2-13 的部分乙酰化 COS。这种 -壳聚糖酶在制备具有不同 DPs 的壳二糖和部分乙酰化 COS 方面具有很大的潜力。
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