Biochemical and structural properties of zebrafish Capsulin produced by Escherichia coli

Protein Expression and Purification
Chi-Yuan ChouYau-Hung Chen

Abstract

Capsulin is one of the transcription factors involved in regulating cell differentiation but its biochemical properties and structural characteristics are still unclear. In the present study, we cloned capsulin from zebrafish, which produces large numbers of transparent embryos and has well-characterized developmental stages. By alignment, the deduced amino acid sequence of zebrafish Capsulin, which contains a putative bHLH motif, shares very high homology to that of other species with an 72-82% identity. Zebrafish Capsulin was also targeted to the nucleus of mammalian cells when overexpressed by transient transfection. In order to characterize the structural and biochemical properties of zebrafish Capsulin, a recombinant zebrafish Capsulin protein was expressed and purified in Escherichia coli. By circular dichroism spectroscopy, Capsulin was shown to be 55% α-helical. The size distribution assay by analytical ultracentrifugation indicated that it existed as a monomer-dimer mixture. The results suggested that the recombinant Capsulin has a well-organized and functional structure. Finally, endogenous Capsulin was distributed mainly in the epicardial cells of zebrafish by immunohistochemistry analysis using antibodies raised aga...Continue Reading

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Citations

Jun 1, 2013·Journal of Toxicologic Pathology·I-Ting TsaiYun-Hsin Wang

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