High-Level Production of Human Leptin by Fed-Batch Cultivation of Recombinant Escherichia coli and Its Purification
AUTOR(ES)
Jeong, Ki Jun
FONTE
American Society for Microbiology
RESUMO
Human leptin is a 16-kDa (146-amino-acid) protein that is secreted from adipocytes and influences body weight homeostasis. In order to obtain high-level production of leptin, the human obese gene coding for leptin was expressed in Escherichia coli BL21(DE3) under the strong inducible T7 promoter. The recombinant leptin was produced as inclusion bodies in E. coli, and the recombinant leptin content was as high as 54% of the total protein content. For production of recombinant human leptin in large amounts, pH-stat fed-batch cultures were grown. Expression of leptin was induced at three different cell optical densities at 600 nm (OD600), 30, 90, and 140. When cells were induced at an OD600 of 90, the amount of leptin produced was 9.7 g/liter (37% of the total protein). After simple purification steps consisting of inclusion body isolation, denaturation and refolding, and anion-exchange chromatography, 144.9 mg of leptin that was more than 90% pure was obtained from a 50-ml culture, and the recovery yield was 41.1%.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=91452Documentos Relacionados
- High-Level Production of Poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate) by Fed-Batch Culture of Recombinant Escherichia coli
- Production of poly(3-hydroxybutyrate) by fed-batch culture of filamentation-suppressed recombinant Escherichia coli.
- Recombinant Erwinia carotovora l-asparaginase II production in Escherichia coli fed-batch cultures
- Production of Poly(3-Hydroxybutyrate) by Fed-Batch Culture of Recombinant Escherichia coli with a Highly Concentrated Whey Solution
- SIMULTANEOUS LIPID AND CAROTENOID PRODUCTION BY STEPWISE FED-BATCH CULTIVATION OF Rhodotorula mucilaginosa WITH CRUDE GLYCEROL