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Improved purification, crystallization and crystallographic study of Hyd‐2‐type [NiFe]‐hydrogenase from Citrobacter sp. S‐77
Author(s) -
Muhd Noor Noor Dina,
Nishikawa Koji,
Nishihara Hirofumi,
Yoon KiSeok,
Ogo Seiji,
Higuchi Yoshiki
Publication year - 2016
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x15024152
Subject(s) - trypsin , crystallization , polyethylene glycol , chemistry , resolution (logic) , citrobacter , crystallography , nuclear chemistry , enzyme , escherichia coli , biochemistry , organic chemistry , enterobacteriaceae , artificial intelligence , computer science , gene
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 P 2 1 , 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 O 2 , indicating that it has clear O 2 tolerance.