中文摘要：

嗜热古菌蛋白Ssh10b突变体[P62A]Ssh10b具有良好的热稳定性，[P62A]Ssh10b的结构测定结果显示，其以螺旋上残基K48和D51之间形成了一对盐键，而且，突变D51将影响蛋白的热稳定性．为了探索DS1的突变对蛋白热稳定性的影响，构建了突变体[D51N／P62A]Ssh10b的质粒，并获得了高纯度的15N和13C双标记[D51N／P62A]Ssh10b．通过对异核三共振NMR实验数据的解析，完成了对[D51N／P62A]Ssh10b的主链共振近乎完全的指认．比较突变以及未突变蛋白质的主链1H^N和15N化学位移，在[P62A]Ssh10b的结构基础上进行分析发现，D51N突变显著地影响了以螺旋骨架构象，并进一步影响到古菌Lβ2α2loop区域、胆的N端区域、Lβ3β4loop的C端区域以及β3与Lβ3β4loop的交界区域．结果表明，由于D51N突变破坏了α2螺旋上K48和D51之间的盐键，影响了[D51N／P62A]Ssh10b的α2螺旋构象，并影响到蛋白的其它相关部位的局部构象，说明[P62A]Ssh10b的高热稳定性可能与其溶液构象密切相关．为进一步运用NMR研究[D51N／P62A]Ssh10b分子结构特性与耐热机制间的关系奠定了基础．

英文摘要：

The P62A mutant hyperthermophilic archaeal protein Ssh10b, [ P62A] Ssh10b, is highly thermostable. The structure determination shows that a salt-bridge K48-D51 is formed in helix α2, and mutation of D51 can influence the thermostability of protein. In order to understand the effect of mutation on the stability of protein, we constructed the D51 N-mutant [ P62A ] Ssh10b plasmid, and obtained the 15N and 13C double-labeled [ D51N/P62A] Ssh10b protein with a high purity. The experimental data of triple-resonance NMR experiments provided nearly complete backbone resonance assignments. Comparing the main chain 1H^N and 15N chemical shifts for [ P62A] Ssh10b with its mutant variant and analyzing the chemical shift differences on the basis of [ P62A] Ssh10b structure, it was found that D51N mutation influenced the backbone conformation of helix α2. The further effect was observed in the structural region consisting of Loop Lβ2α2, the N-terminal of strandβ4, and the C-terminal of loop Lβ3β4, and in the region around loop Lβ3β4 as well. This revealed that D51 N mutation disrupted the salt bridge of K48-D51 on helix α2 and influenced not only the backbone con-formation of helix α2 but also the local conformations of other structural regions in the protein. Therefore, the high thermostability of [ P62A] Ssh10b may be correlated closely to its tertiary conformation.