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Int. J. Mol. Sci. 2009, 10(1), 28-36; doi:10.3390/ijms10010028
Article

Over-expression, Rapid Preparation and Some Properties of C-terminal BARc Region in PICK1

1, 2,#,* , 2,#, 2,* , 2 and 3
Received: 19 October 2008; in revised form: 16 December 2008 / Accepted: 18 December 2008 / Published: 27 December 2008
(This article belongs to the Section Biochemistry, Molecular Biology and Biophysics)
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Abstract: A DNA fragment encoding C-terminal BARc region (amino acids 128-416) of rat PICK1 (NP_445912 ) was inserted into a modified vector pMAL-s involving human rhinovirus 3C protease cleavage site to produce a recombinant plasmid, pMAL-s-barc. The construct can express the fusion protein, MBP-BARc in the soluble form in E.coli. To remove the MBP tag, MBP-BARc purified from amylose beads was digested with human rhinovirus 3C protease and the cleavage efficiency is about 95% when the ratio of protein / enzyme (w/w) reaches 50:1, as analyzed on SDS-PAGE. The enzymatic reaction mixture was rapidly separated into two parts, MBP in the supernatant and BARc in the precipitate at the concentration of 1 M ammonium sulfate. In such case, the target protein BARc could be economically produced in a soluble state to be as the sample for measuring its biochemical function, for example, protein-protein interaction and protein-lipid combination.
Keywords: PICK1; BARc; human rhinovirus 3C protease cleavage; ammonium sulfate precipitation; inter-molecular interaction PICK1; BARc; human rhinovirus 3C protease cleavage; ammonium sulfate precipitation; inter-molecular interaction
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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MDPI and ACS Style

Xiao, H.; Shi, Y.; Yuan, J.; Huang, Y.; Wang, J. Over-expression, Rapid Preparation and Some Properties of C-terminal BARc Region in PICK1. Int. J. Mol. Sci. 2009, 10, 28-36.

AMA Style

Xiao H, Shi Y, Yuan J, Huang Y, Wang J. Over-expression, Rapid Preparation and Some Properties of C-terminal BARc Region in PICK1. International Journal of Molecular Sciences. 2009; 10(1):28-36.

Chicago/Turabian Style

Xiao, Hong; Shi, Yawei; Yuan, Jingming; Huang, Yuming; Wang, Junhua. 2009. "Over-expression, Rapid Preparation and Some Properties of C-terminal BARc Region in PICK1." Int. J. Mol. Sci. 10, no. 1: 28-36.



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