PicW2 from Picea wilsonii: preparation, purification, crystallization and X-ray diffraction analysis.

ABSTRACT: 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 C121, with unit-cell parameters a = 121.55, b = 33.26, c = 73.39?Å, ? = ? = 90.00, ? = 109.01°. The asymmetric unit contained one molecule of the protein, with a corresponding Matthews coefficient of 2.87?Å3?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.

SUBMITTER: Zhang B 

PROVIDER: S-EPMC5987745 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature