Key Laboratory of Glycoscience & Glycotechnology of Shandong Province, Key Laboratory of Marine Drugs, Chinese Ministry of Education, Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, School of Medicine and Pharmacy, Ocean University of China, 5 Yushan Road, Qingdao 266003, China.
Key Laboratory of Glycoscience & Glycotechnology of Shandong Province, Key Laboratory of Marine Drugs, Chinese Ministry of Education, Laboratory for Marine Drugs and Bioproducts, Qingdao National Laboratory for Marine Science and Technology, School of Medicine and Pharmacy, Ocean University of China, 5 Yushan Road, Qingdao 266003, China.
Int J Biol Macromol. 2018 Apr 1;109:880-887. doi: 10.1016/j.ijbiomac.2017.11.071. Epub 2017 Nov 16.
CsnA, a chitosanase from Renibacterium sp. QD1, has great potential for industrial applications due to its high yield and broad pH stability. In this study, a specific Glu160 in CsnA was identified by sequence alignment, and structural analysis and MD simulation predicted that Glu160 formed a hydrogen-bond network with Lys163 and Thr114. To evaluate the effect of the network, we constructed four mutants, including E160A, E160Q, K163A, and T114A, which partially or completely destroy this network. Characterization of these mutants demonstrated that the disruption of the network significantly decreased the enzyme thermostability. The underlying mechanisms responsible for the change of thermostability analyzed by circular dichroism spectroscopy revealed that the hydrogen-bond network conferred the structural stability of CsnA. Moreover, the length of the side chain of residue at 160 impacted conformational stability of the enzyme. Taken together, the hydrogen-bond network around Glu160 plays important roles in stabilization of CsnA.
壳聚糖酶 CsnA 是从海洋细菌 Renibacterium sp. QD1 中分离得到的，具有产量高、pH 稳定性广等优点，具有很大的工业应用潜力。本研究通过序列比对、结构分析和 MD 模拟预测，鉴定出 CsnA 中的一个特定Glu160，该Glu160 与 Lys163 和 Thr114 形成氢键网络。为了评估该网络的作用，我们构建了四个突变体，包括 E160A、E160Q、K163A 和 T114A，它们部分或完全破坏了这个网络。对这些突变体的表征表明，该网络的破坏显著降低了酶的热稳定性。圆二色光谱分析揭示了稳定性变化的潜在机制，即氢键网络赋予了 CsnA 结构稳定性。此外，残基 160 侧链的长度影响了酶的构象稳定性。综上所述，Glu160 周围的氢键网络在 CsnA 的稳定中发挥着重要作用。
