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Purification, crystallization and preliminary X-ray crystallographic analysis of the CIDE-N domain of Fsp27.


ABSTRACT: Fsp27, a member of the CIDE protein family which is selectively expressed in adipocytes, has emerged as a novel regulator for unilocular lipid droplet (LD) formation, lipid metabolism, differentiation of adipocytes and insulin sensitivity. An LD is a subcellular compartment that is used by adipocytes for the efficient storage of fats. The CIDE-N domain of Fsp27 functions as a recruitment platform that induces the correct configuration of the Fsp27 CIDE-C domain to facilitate LD fusion. This study reports the high-yield expression of the mouse Fsp27 CIDE-N domain in Escherichia coli; a two-step purification protocol with high efficiency was established and crystallographic analysis was performed. The purity of the recombinant Fsp27 was >95% as assessed by SDS-PAGE. Crystals were obtained at 291?K using 28% polyethylene glycol 4000 as a precipitant. Diffraction data were collected to 1.92?Å resolution and the crystal belonged to space group P6(5), with unit-cell parameters a=b=63.3, c=37.4?Å, ?=?=90, ?=120°. The components of the crystal were identified by ion-trap LC/MS/MS spectrometric analysis. The structure has been solved by molecular replacement and refinement is in progress.

SUBMITTER: Wang X 

PROVIDER: S-EPMC3509981 | biostudies-literature | 2012 Dec

REPOSITORIES: biostudies-literature

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Purification, crystallization and preliminary X-ray crystallographic analysis of the CIDE-N domain of Fsp27.

Wang Xiaodan X   Zhang Bo B   Xu Duo D   Gao Jinlan J   Wang Linfang L   Wang Zhi Z   Shan Yaming Y   Yu Xianghui X  

Acta crystallographica. Section F, Structural biology and crystallization communications 20121114 Pt 12


Fsp27, a member of the CIDE protein family which is selectively expressed in adipocytes, has emerged as a novel regulator for unilocular lipid droplet (LD) formation, lipid metabolism, differentiation of adipocytes and insulin sensitivity. An LD is a subcellular compartment that is used by adipocytes for the efficient storage of fats. The CIDE-N domain of Fsp27 functions as a recruitment platform that induces the correct configuration of the Fsp27 CIDE-C domain to facilitate LD fusion. This stud  ...[more]

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