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http://dx.doi.org/10.6564/JKMRS.2010.14.2.105

Expression, Purification and NMR studies of SH3YL1 SH3 domain  

Shrestha, Pravesh (Structural Biochemistry and Molecular Biophysics Laboratory, Department of Biochemistry, College of Life Science & Biotechnology, Yonsei University)
Yun, Ji-Hye (Structural Biochemistry and Molecular Biophysics Laboratory, Department of Biochemistry, College of Life Science & Biotechnology, Yonsei University)
Lee, Weon-Tae (Structural Biochemistry and Molecular Biophysics Laboratory, Department of Biochemistry, College of Life Science & Biotechnology, Yonsei University)
Publication Information
Journal of the Korean Magnetic Resonance Society / v.14, no.2, 2010 , pp. 105-116 More about this Journal
Abstract
SH3YL1, a novel protein containing one Src homology 3 domain at the carboxyl terminus was first detected in mouse anagen skin cDNA. This protein had a significant homology with YHRO 16c/Ysc 84, the yeast Src homology 3 domain-containing protein. The sequence identity was remarkable at the carboxyl and amino-terminal Src homology 3 domain, suggesting that the novel protein is a mouse homolog of the yeast protein and thus was termed as SH3YL1. SH3YL1 is composed of two domains, a DUF500 at N-termini and a SH3 domain at C-termini. In our study we cloned the SH3 domain in bacterial expression system in Escherichia coli using pET32a vector with TEV protease cleavage site and purified as a monomer using affinity chromatography. The N-terminal poly-Histidine tag was cleaved with TEV protease and target protein was used for backbone studies. Our study showed that SH3 domain primarily consists of $\beta$-sheet which is in consistence with previous result performed on the truncated SH3 domain of SH3YL1.
Keywords
Src Homology 3 domain; DUF500 domain; NMR spectroscopy; TALOS analysis; Cloning; Purification;
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