Expression, purification and characterization of soluble recombinant periostin protein produced by Escherichia coli.
Periostin is a matricellular protein participating in the tissue remodelling of damaged cardiac tissue after acute myocardial infarction and of the periodontal ligament in mice. However, further studies on the periostin protein have been limited by the intrinsic difficulty of purifying this protein produced in Escherichia coli due to its insolubility. Here, we demonstrate the expression of recombinant periostin protein with high solubility and monodispersity in E. coli. Periostin is composed of an amino-terminal EMI domain, a tandem repeat of 4 fas1 domains (RD1-4), and a carboxyl-terminal region (CTR). We expressed the RD4-CTR region tagged with GST at amino-terminal and 6x Histidine at carboxyl-terminal end in E. coli. The recombinant protein was purified by using GSH-Sepharose and nickel chelation affinity chromatography, followed by gel filtration chromatography. The RD4-CTR protein exhibited high solubility and monodispersity. On average, 9.1 mg of purified RD4-CTR was routinely obtained from 1 L of culture media. Furthermore, the RD4-CTR was biochemically active, because it bound to the RD1-4, the same as intact periostin protein that had been purified from mammalian cells. Our results should enable us to produce the periostin recombinant protein in large quantities and facilitate future studies on functional and structural analyses of periostin.
Takayama
Issei
I
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Midori-ku, Nagatsuta, Yokohama 226-8501, Japan.
Kii
Isao
I
Kudo
Akira
A
D017398:Alternative Splicing / Q000235:genetics; D000818:Animals; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D051379:Mice; D050505:Mutant Proteins / Q000378:metabolism; D010513:Periodontal Ligament / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D012995:Solubility
D015815:Cell Adhesion Molecules; D050505:Mutant Proteins; C082838:Postn protein, mouse; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins