Abstract
Escherichia coli has been most extensively used for the large-scale production of therapeutic proteins, which do not require complex glycosylation for bioactivity. In recent years tremendous progress has been made on the molecular biology, fermentation process development and protein refolding from inclusion bodies for efficient production of therapeutic proteins using E. coli. High cell density fermentation and high throughput purification of the recombinant protein from inclusion bodies of E. coli are the two major bottle necks for the cost effective production of therapeutic proteins. The aim of this review is to summarize the developments both in high cell density, high productive fermentation and inclusion body protein refolding processes using E. coli as an expression system. The first section deals with the problems of high cell density fermentation with an aim to high volumetric productivity of recombinant protein. Process engineering parameters during the expression of ovine growth hormone as inclusion body in E. coli were analyzed. Ovine growth hormone yield was improved from 60 mg L-1 to 3.2 g L-1 using fed-batch culture. Similar high volumetric yields were also achieved for human growth hormone and for recombinant bonnet monkey zona pellucida glycoprotein expressed as inclusion bodies in E. coli. The second section deals with purification and refolding of recombinant proteins from the inclusion bodies of E. coli. The nature of inclusion body protein, its characterization and isolation from E. coli has been discussed in detail. Different solubilization and refolding methods, which have been used to recover bioactive protein from inclusion bodies of E. coli have also been discussed. A novel inclusion body protein solubilization method, while retaining the existing native-like secondary structure of the protein and its subsequent refolding in to bioactive form, has been discussed. This inclusion body solubilization and refolding method has been applied to recover bioactive recombinant ovine growth hormone, recombinant human growth hormone and bonnet monkey zona pellucida glycoprotein from the inclusion bodies of E. coli.
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Panda, A.K. (2003). Bioprocessing of Therapeutic Proteins from the Inclusion Bodies of Escherichia coli . In: Ghose, T.K., et al. Biotechnology in India II. Advances in Biochemical Engineering/Biotechnology, vol 85. Springer, Berlin, Heidelberg. https://doi.org/10.1007/3-540-36466-8_3
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