Production of interferon-a in high cell density cultures of recombinant Escherichia coli and its single step puri®cation from refolded inclusion body proteins
- نوع فایل : کتاب
- زبان : انگلیسی
- مؤلف : K. R. Babu ل S. Swaminathan ل S. Marten N. Khanna ل U. Rinas
- چاپ و سال / کشور: 2000
Description
Escherichia coli TG1 transformed with a temperature-regulated interferon-a expression vector was grown to high cell density in deRned medium con- taining glucose as the sole carbon and energy source, utilizing a simple fed-batch process. Feeding was carried out to achieve an exponential increase in biomass at growth rates which minimized acetate production. Thermal induction of such high cell density cultures resulted in the production of 4 g interferon-a/l culture broth. Interferon-a was produced exclusively in the form of insoluble inclusion bodies and was solubilized under denaturing conditions, refolded in the presence of argi- nine and puriRed to near homogeneity, utilizing single- step ion-exchange chromatography on Q-Sepharose. The yield of puriRed interferon-a was 300 mg/l with respect to the original high cell density culture broth (overall yield of 7.5% active interferon-a). The puriRed recombinant interferon-a was found by di€erent criteria to be predominantly monomeric and possessed a speciRc bioactivity of 2.5 پL 108 IU/mg based on viral cytopathic assay.
Appl Microbiol Biotechnol (2000) 53: 655-660 Received: 8 October 1999 / Received revision: 8 December 1999 / Accepted: 12 December 1999
