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PicW2 from Picea wilsonii : preparation, purification, crystallization and X‐ray diffraction analysis
Author(s) -
Zhang Bei,
Guo Gangxing,
Lu Fang,
Song Ying,
Liu Yong,
Xu JiChen,
Gao Wei
Publication year - 2018
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/s2053230x18007537
Subject(s) - crystallization , protein crystallization , chemistry , crystal (programming language) , size exclusion chromatography , crystal structure , crystallography , x ray crystallography , molecular replacement , chromatography , diffraction , organic chemistry , enzyme , physics , optics , computer science , programming language
Low temperature is a major limiting factor for plant growth and development. Dehydrin proteins are generally induced in response to low‐temperature stress. In previous research, a full‐length dehydrin gene, PicW2 , was isolated from Picea wilsonii and its expression was associated with hardiness to cold. In order to gain insight into the mechanism of low‐temperature tolerance by studying its three‐dimensional crystal structure, prokaryotically expressed PicW2 dehydrin protein was purified using chitosan‐affinity chromatography and gel filtration, and crystallized using the vapour‐diffusion method. The crystal grew in a condition consisting of 0.1 M HEPES pH 8.0, 25%( w / v ) PEG 3350 using 4 mg ml −1 protein solution at 289 K. X‐ray diffraction data were collected from a crystal at 100 K to 2.82 Å resolution. The crystal belonged to space group C 121, with unit‐cell parameters a = 121.55, b = 33.26, c = 73.39 Å, α = γ = 90.00, β = 109.01°. The asymmetric unit contained one molecule of the protein, with a corresponding Matthews coefficient of 2.87 Å 3 Da −1 and a solvent content of 57.20%. Owing to a lack of structures of homologous dehydrin proteins, molecular‐replacement trials failed. Data collection for selenium derivatization of PicW2 and crystal structure determination is currently in progress.