C端柔性区域对多糖降解菌来源的麦芽五糖生成酶冷适性的调控
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Q556.2

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Modulation of Cold Adaptation of Saccharophagus degradans Maltopentaose-Forming Amylase via Its C-Terminal Flexible Region
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    摘要:

    冷适麦芽五糖生成酶在低温下具有较高的催化活力,同时能够在室温下快速水解淀粉并特异性地生成具有营养功效的麦芽五糖,因此在食品、医药等领域具有广阔的应用前景。作者以来源于Saccharophagus degradans的两种麦芽五糖生成酶(SdG5A和SdG5A-CD)为研究对象,将其分别表达于Bacillus subtilis中,并分析比较了重组酶的冷适性。结果表明,重组SdG5A在0 ℃下可以保持27.8%的酶活力,在室温下生产麦芽五糖的得率高达48.6%,具有较强的冷适性;而不含有linker和淀粉结合域(SBD)的SdG5A-CD不具有冷适性。为了探究SdG5A的冷适性机理,利用RoseTTAFold构建了结构模型,并利用分子动力学模拟等方法分析了结构柔性。结果表明,位于SdG5A C端的linker-SBD区域具有极高的柔性,且在0 ℃下的均方根涨落与45 ℃保持一致,说明linker-SBD结构高柔性的分子特征是影响SdG5A冷适性的关键因素。

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    Cold-adapted maltopentaose-forming amylases have relatively high catalytic activities at low temperatures and enable an efficient and specific conversion of starch to functional maltopentaose at room temperature, resulting in a broad applications in food, medical and pharmaceutical fields. In this study, SdG5A and SdG5A-CD from Saccharophagus degradans were expressed in Bacillus subtilis. Further, the cold adaptation of the recombinant enzymes was characterized and compared. The results showed that recombinant SdG5A retained 27.8% of its maximal activity at 0 ℃ and could produce 48.6% of maltopentaose at room temperature, indicating a strong cold adaptation of SdG5A. In contrast, SdG5A-CD, lacking the linker and starch-binding domain (SBD), did not show any cold adaptation. To investigate the mechanisms that might underlie the cold adaptation of SdG5A, the structure was predicted by RoseTTAFold and the structural flexibility was analyzed through molecular dynamics simulation. An extremely high flexibility in the C-terminal linker-SBD region was observed and its root-mean-square fluctuation at 0 ℃ was consistent with that at 45 ℃, indicating that the highly flexible linker-SBD region acted as a critical component for the cold adaptation of SdG5A.

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丁宁,李才明,班宵逢,顾正彪,李兆丰. C端柔性区域对多糖降解菌来源的麦芽五糖生成酶冷适性的调控[J].食品与生物技术学报,2022,41(10):67-76.

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  • 在线发布日期: 2022-11-16
  • 出版日期: 2022-10-25

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