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Expression, Purification, and Secondary Structure Characterization of Recombinant KCTD1

Fanghua Mei1,2#, Jin Xiang3#, Song Han2, Yuan He2, Yajing Lu2, Jian Xu2, Deyin Guo2, Gengfu Xiao1, Po Tien1, and Guihong Sun2*

1The State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China

2School of Basic Medical Sciences, Wuhan University, Wuhan 430071, China; fax: 86-27-6875-9142; E-mail: ghsunlab@whu.edu.cn

3College of Pharmacy, Wuhan University, Wuhan 430072, China

# These authors contributed equally to this work.

* To whom correspondence should be addressed.

Received March 29, 2012; Revision received May 18, 2012
Potassium channel tetramerization domain containing 1 (KCTD1) contains a BTB domain, which can facilitate protein–protein interactions that may be involved in the regulation of signaling pathways. Here we describe an expression and purification system that can provide a significant amount of recombinant KCTD1 from Escherichia coli. The cDNA encoding human KCTD1 was amplified and cloned into the expression vector pET-30a(+). The recombinant protein was expressed in E. coli BL21(DE3) cells and subsequently purified using affinity chromatography. To confirm that KCTD1 was correctly expressed and folded, the molecular weight and conformation were analyzed using mass spectroscopy, Western blot, and circular dichroism. Optimizing KCTD1 expression and investigating its secondary structure will provide valuable information for future structural and functional studies of KCTD1 and KCTD family proteins.
KEY WORDS: KCTD1, circular dichroism spectroscopy, secondary structure

DOI: 10.1134/S0006297912080160