[Expression, purification and characterization of human prorelaxin-like-protein H2 in Escherichia coli].
The cDNA gene encoding human prorelaxin H2 was subcloned from plasmid pMAL p2X-hRLX H2 into the EcoRI and BamHI sites of prokaryotic expression vector pBV220, resulting in the recombinant plasmid pBV220-hPRLX H2, which was subsequently transformed into Escherichia coli DH5arg;. The target gene was highly expressed in the form of inclusion body by thermoinduction at 42 degrees. Recombinant human Met-prorelaxin-like-protein H2 was refolded in vitro, purified by Sephadex G-75 gel filtration chromatography and reverse phase fast protein liquid chromatography. The yield was approximately 2-3 mg/L. The purified recombinant human Met-prorelaxin-like-protein H2 was shown to be a single band by 15% SDS-PAGE and gave correct amino acid composition of Met-prorelaxin. Molecular weight of the purified recombinant human Met-prorelaxin-like-protein H2 was measured to be 18 390.4 (calculated theoretical value 18 392.3) by matrix assisted laser desorption ionization time-of-flight mass spectroscopy.
Chen
Li-Ming
LM
National Laboratory of Protein Engineering and Plant Genetic Engineering, Peking University, Beijing 100871, China. lxzyx@pku.edu.cn
Yang
Xing-Wen
XW
Tang
Jian-Guo
JG
D000596:Amino Acids / Q000032:analysis; D004926:Escherichia coli; D015870:Gene Expression; D005818:Genetic Engineering; D005822:Genetic Vectors; D006801:Humans; D011498:Protein Precursors / Q000032:analysis / Q000235:genetics*; D012065:Relaxin / Q000032:analysis / Q000235:genetics*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D000596:Amino Acids; D011498:Protein Precursors; C095068:RLN2 protein, human; D012065:Relaxin