A thermostable and alkaline GDSL-motif esterase from Bacillus sp. K91: crystallization and X-ray crystallographic analysis.
A thermostable and alkaline GDSL-motif esterase from Bacillus sp. K91: crystallization and X-ray crystallographic analysis.
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来自芽孢杆菌 K91 的热稳定性和碱性 GDSL 基序酯酶：结晶和 X 射线晶体分析
DOI：
10.1107/s2053230x18000353
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发表时间：
2018-02-01
期刊：
影响因子：
--
通讯作者：
Meng Z
中科院分区：
文献类型：
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作者：
Ding J;Zhu H;Ye Y;Li J;Han N;Wu Q;Huang Z;Meng Z
采用纯化、结晶和x射线晶体学分析的方法对芽孢杆菌K91中gdsl基序酯酶Est8的催化机制进行了研究。来自嗜热细菌Bacillus sp. K91的酯酶Est8属于GDSL家族，对多种乙酰化化合物有活性，包括7-氨基头孢孢酸。与GDSL家族的其他酯酶相比，催化残基Asp182和His185比Ser11残基对Est8的催化活性更关键。为了更好地了解Est8的生化和酶学性质，我们对重组Est8蛋白进行了纯化和结晶。以2.0 M硫酸铵、5%(v/v) 2-丙醇为结晶液，采用悬垂气相扩散法制备了Est8结晶。x射线衍射数据采集到分辨率为2.30 Å， R归并率为16.4%，晶体属于空间群P41212或P43212，单胞参数a = b = 68.50, c = 79.57 Å。
Purification, crystallization and X-ray crystallographic analysis were employed to determine the catalytic mechanism of Est8, a GDSL-motif esterase from Bacillus sp. K91. The esterase Est8 from the thermophilic bacterium Bacillus sp. K91 belongs to the GDSL family and is active on a variety of acetylated compounds, including 7-aminocephalosporanic acid. In contrast to other esterases of the GDSL family, the catalytic residues Asp182 and His185 were more pivotal for the catalytic activity of Est8 than the Ser11 residue. To better understand the biochemical and enzymatic properties of Est8, recombinant Est8 protein was purified and crystallized. Crystals of Est8 were obtained by the hanging-drop vapour-diffusion method using 2.0 M ammonium sulfate, 5%(v/v) 2-propanol as the crystallization solution. X-ray diffraction data were collected to a resolution of 2.30 Å with an R merge of 16.4% from a crystal belonging to space group P41212 or P43212, with unit-cell parameters a = b = 68.50, c = 79.57 Å.