[关键词]
[摘要]
为了解酪氨酸酚裂解酶(TPL)热稳定性与序列之间的关系,为突变产生耐热性的酪氨酸酚裂解酶提供参考,利用生物信息学对来源于Symbiobacterium toebii的耐热酪氨酸酚裂解酶StTPL、来源于Fusobacterium nucleatum的耐热酪氨酸酚裂解酶FnTPL和来源于Citrobacter freundii的中温酪氨酸酚裂解酶CfTPL进行进化树、一级结构、二级结构和三级结构的比较。随着TPL中一级结构中脯氨酸含量增多,最适反应温度的增加,StTPL、FnTPL和CfTPL的脯氨酸含量分别为5.02%、3.70%、2.85%,二级结构中β-折叠和转角较多,分别为25.51%、25.00%、23.91%,这些特征符合文献报道的耐热酶和中温酶的区别。将StTPL、FnTPL和CfTPL的三维结构叠加比对发现,310-313残基区域、残基13、82-88残基区域叠加效果差,313残基突变有利于CfTPL提高热稳定性,StTPL中的Ala13、Glu83、Thr407突变有利于StTPL热稳定性,推测以上残基区域和热稳定性相关,也符合已有文献报道的情况。此外,有关生物信息学分析为进一步理解超稳酶的超稳机制以及筛选改造新的超稳酶提供了初步参考。
[Key word]
[Abstract]
In order to understand the relationship between the thermostability of tyrosine phenol lyase (TPL) and its sequence, and to provide a reference for mutating the tyrosine phenol lyase to produce thermotolerant tyrosine phenol lyase, bioinformatics was used to analyze the thermostable tyrosine phenollyases, StTPL derived from Symbiobacterium toebii, FnTPL derived from Fusobacterium nucleatum, and mesophilic CfTPL derived from Citrobacter freundii. These lyases were compared in terms of phylogenetic tree, primary structure, secondary structure and tertiary structure. With the increase of the proline content in the primary structure of TPL, the optimal reaction temperature increased. The proline contents of StTPL, FnTPL and CfTPL were 5.02%, 3.70% and 2.85%, respectively. There were more β-sheets and turns in the secondary structure, which were 25.51%, 25.00%, and 23.91%, respectively. These characteristics were in line with the difference between thermostable enzymes and mesophilic enzymes reported in the literature. The comparison of the three-dimensional structures of StTPL, FnTPL and CfTPL revealed that the 310-313 residue region, residue 13, and 82-88 residue region had a poor superposition effect. The mutation of the 313 residue was conducive to the thermalstability of CfTPL. The mutation ofAla13, Glu83 and Thr407 residue was conducive to the thermalstability of StTPL. It is speculated that the above residue regions are related to thermalstability, which is also in line with the findings reported in the literature. In addition, the relevant bioinformatics analysis provides a preliminary reference for further understanding the hyperstability mechanism of hyperstable enzymes and screening and transforming new hyperstable enzymes.
[中图分类号]
[基金项目]
国家重点研发计划项目(2018YFA0901700)