Production of recombinant mink growth hormone in E. coli.
Escherichia coli cells expressing mink (Mustela vison) growth hormone were grown in a batch fermentation process. The expression level was estimated to be 27% of the total cellular protein after 3 h of induction with 1 mM isopropyl beta-D-thiogalactoside (IPTG). If the expression of mink growth hormone (mGH) was induced with 0.2 mM IPTG, the concentration of target protein was slightly lower and was found to be 23% at the same time after induction. mGH expressed as inclusion bodies was solubilized in 8 M urea and renatured by dilution protocol at a protein concentration of 1.4-2.1 mg/ml in the presence of glutathione pair in a final concentration of 11.3 mM. [GSH]/[GSSG] ratio equal to 2/1 was used. Two-step purification process comprising of ion-exchange chromatography on Q-Sepharose and hydrophobic chromatography on Phenyl-Sepharose was developed. Some 25-30 mg of highly purified and biologically active mGH was obtained from 4 g of biomass. The method presented in this study allows producing large quantities of mGH and considering initiation of scientific investigation on mGH effect on mink in vivo and availability in fur industry.
Sereikaite
Jolanta
J
Department of Chemistry and Bioengineering, Faculty of Fundamental Sciences, Vilnius Gediminas Technical University, Sauletekio al. 11, 2040 Vilnius, Lithuania. sjolanta@fm.vtu.lt
Statkute
Alina
A
Morkunas
Mindaugas
M
Radzevicius
Kostas
K
Borromeo
Vitaliano
V
Secchi
Camillo
C
Bumelis
Vladas-Algirdas
VA
D000818:Animals; D001709:Biotechnology / Q000379:methods*; D002460:Cell Line; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D015964:Gene Expression Regulation, Bacterial; D013006:Growth Hormone / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008907:Mink / Q000378:metabolism*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D003470:Culture Media; D011994:Recombinant Proteins; D013006:Growth Hormone