Improved purification, crystallization and crystallographic study of Hyd-2 type [NiFe] hydrogenase from Citrobacter sp. S-77
Improved purification, crystallization and crystallographic study of Hyd-2 type [NiFe] hydrogenase from Citrobacter sp. S-77
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改进了来自柠檬酸杆菌属的 Hyd-2 型 [NiFe] 氢化酶的纯化、结晶和晶体学研究。
DOI：
10.1107/s2053230x15024152
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发表时间：
2016
期刊：
影响因子：
--
通讯作者：
Yoshiki
中科院分区：
文献类型：
--
作者：
Muhd Noor;N.D.; Nishikawa;Koji; Nishihara;Hirofumi; Yoon;Ki-Seok; Ogo;Seiji; Higuchi;Yoshiki
柠檬酸杆菌hyd-2型[NiFe]-氢酶的纯化工艺。通过对S-77进行层析前胰酶处理，对其进行了改良。用聚乙二醇作沉淀剂，用坐滴气相扩散法成功地结晶了胰酶处理和未处理的纯化蛋白质样品。经胰酶处理的蛋白质晶胞参数为a=63.90，b=118.89，c=96.70 ä，β=100.61°；未经胰酶处理的样品，晶胞参数为a=65.38，b=121.45，c=98.63 ä，β=102.29°。从胰酶处理的蛋白质得到的晶体衍射率为1.60 ä，这比没有胰酶处理的2.00 ä的分辨率要好得多。柠檬酸杆菌的[NiFe]-氢酶。S-77在含有一定量O2的情况下仍保持催化活性，表明其具有明显的耐氧性。
The purification procedure of Hyd-2-type [NiFe]-hydrogenase from Citrobacter sp. S-77 was improved by applying treatment with trypsin before chromatography. Purified protein samples both with and without trypsin treatment were successfully crystallized using the sitting-drop vapour-diffusion method with polyethylene glycol as a precipitant. Both crystals belonged to space group P21, with unit-cell parameters a = 63.90, b = 118.89, c = 96.70 Å, β = 100.61° for the protein subjected to trypsin treatment and a = 65.38, b = 121.45, c = 98.63 Å, β = 102.29° for the sample not treated with trypsin. The crystal obtained from the trypsin-treated protein diffracted to 1.60 Å resolution, which is considerably better than the 2.00 Å resolution obtained without trypsin treatment. The [NiFe]-hydrogenase from Citrobacter sp. S-77 retained catalytic activity with some amount of O2, indicating that it has clear O2 tolerance.