Studies on hyperthermostable aldolase : characteristics, structure and application

超热稳定醛缩酶的研究：特性、结构和应用

基本信息

批准号：
15560677
负责人：
SAKURABA Haruhiko
金额：
$ 2.37万
依托单位：
The University of Tokushima
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

A gene encoding a D-2-deoxyribose-5-phosphate aldolase (DERA) homologue was identified in the hyperthermophilic archaeon Aeropyrum pernix. The gene was overexpressed in Escherichia coli, and the produced enzyme was purified and characterized. The enzyme was an extremely thermostable DERA ; the activity was not lost after incubation at 100℃ for 10 min. The enzyme had a molecular mass of about 93 kDa and consisted of four subunits with an identical molecular mass of 24 kDa. This shows the first presence of the tetrameric DERA. The three-dimensional structure of the enzyme was determined by x-ray analysis. The subunit folded into an α/β barrel. The asymmetric unit consisted of two homologous subunits, and a crystallographic 2-fold axis generated the functional tetramer. Compared with the structure of the E. coli DERA, the mainchain coordinate of the monomer of A. pernix enzyme was quite similar to that of the E. coli enzyme. A large difference in hydrophobic interactions and the number of ion pairs was not observed between the monomeric structures of the two enzymes. However, a significant difference in the quaternary structure was observed. The area of the subunit-subunit interface in the dimer of the A. pernix enzyme was much larger than the corresponding one of the E. coli enzyme. In addition, the A. pernix enzyme was 10 amino acids longer than the E. coli enzyme in the N-terminal region and exhibited an additional N-terminal helix. The N-terminal helix produced a unique dimer-dimer interface. This promotes the formation of a functional tetramer of the A. pernix enzyme and strengthens the hydrophobic inter-subunit interactions. These structural features are considered to be responsible for the extremely high stability of the A. pernix enzyme. This is the first description of the structure of hyperthermophilic DERA and of aldolase from the domain of archaea.

在高疗法Aeropyrum Pernix中鉴定了编码D-2-脱氧核糖5-磷酸醛酶（DERA）同源物的基因。该基因在大肠杆菌中过表达，并且产生的酶被纯化和表征。该酶是一种极其温暖的DERA。该活动在100分钟孵育10分钟后不会损失。该酶的分子质量约为93 kDa，由四个亚基组成，其分子量相同24 kDa。这显示了四聚体Dera的第一次存在。通过X射线分析确定酶的三维结构。亚基折叠成α/β桶。不对称单元由两个同源亚基组成，一个晶体学的2倍轴产生了功能性四聚体。与大肠杆菌DERA的结构相比，A. pernix酶的单体的主链坐标与大肠杆菌酶非常相似。在两种酶的单体结构之间未观察到疏水相互作用的巨大差异和离子对的数量。但是，观察到第四纪结构的显着差异。 A. pernix酶二聚体中亚基 - 亚基界面的面积比相应的大肠杆菌酶之一大得多。此外，丙糖曲霉的氨基酸比N末端区域的大肠杆菌长10个氨基酸，并暴露了额外的N末端螺旋。 N末端螺旋产生了独特的二聚体界面。这促进了A. pernix酶的功能四聚体的形成，并强调疏水性增生间相互作用。这些结构特征被认为是A. pernix酶极高稳定性的原因。这是对古细菌域的高疗Dera和醛糖酶结构的第一个描述。

项目成果

期刊论文数量（42）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The First Crystal Structure of Archaeal Aldolase : Unique Tetrameric Structure of 2-De-oxy-D-ribose-5-phosphate Aldolase from Hyperthermophilic Archaea Aeropyrum pernix

古菌醛缩酶的第一个晶体结构：来自超嗜热古菌 Aeropyrum pernix 的 2-脱氧-D-核糖-5-磷酸醛缩酶的独特四聚结构

DOI：
发表时间：
2003
期刊：
J Biol.Chem. 278
影响因子：
0
作者：
H.Sakuraba;H.Sakuraba;Haruhiko Sakuraba;Haruhiko Sakuraba;H.Sakuraba;H.Sakurada;H.Sakuraba;H.Sakuraba
通讯作者：
H.Sakuraba

Novel archaeal Alanine : Glyoxylate aminotransferase from Thermococcus litralis.

新型古细菌丙氨酸：来自柠檬热球菌的乙醛酸转氨酶。

DOI：
发表时间：
2004
期刊：
J Bacteriol. 186
影响因子：
0
作者：
H.Sakuraba
通讯作者：
H.Sakuraba

The First Crystal Structure of Archaeal Aldolase : Unique Tetrameric Structure of 2-Deoxy-D-ribose-5-phosphate Aldolase from Hyperthermophilic Archaea Aeropyrum pernix

古菌醛缩酶的第一个晶体结构：来自超嗜热古菌 Aeropyrum pernix 的 2-脱氧-D-核糖-5-磷酸醛缩酶的独特四聚体结构

DOI：
发表时间：
2003
期刊：
J.Biol.Chem. Vol.278
影响因子：
0
作者：
H.Sakuraba;H.Sakuraba;Haruhiko Sakuraba;Haruhiko Sakuraba;H.Sakuraba;H.Sakurada;H.Sakuraba;H.Sakuraba;H.Sakuraba;T.Ohshima;Haruhiko Sakuraba;Haruhiko Sakuraba
通讯作者：
Haruhiko Sakuraba

H.Sakuraba et al.: "A nicotinamide mononucleotide adenylyltransferase with unique adenylyl group donor specificity from a hyperthermophilic archaeon, Pyrococcus horikoshii OT3"J.Mol.Catalys.B : Enzymatic. 23. 273-279 (2003)

H.Sakuraba 等人：“一种烟酰胺单核苷酸腺苷酸转移酶，具有来自超嗜热古菌 Pyrococcus horikoshii OT3 的独特腺苷酸基供体特异性”J.Mol.Catalys.B：酶促。