Apte-Deshpande Anjali

Publications (2)0 Total impact

• Article: Arabinose Promoter Based Expression of Biologically Active Recombinant Human Growth Hormone in E. coli: Strategies for Over Expression and Simple Purification Methods

  Soorapaneni Sudheerbabu, Apte-Deshpande Anjali, Sabnis-Prasad Ketaki, Kumar Jitendra, Veena A Raiker
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  ABSTRACT: An arabinose promoter based expression system in E. colifor the production and purification of recombinant humangrowth hormone (rhGH) was designed and implemented.The shake flask studies indicated appreciable amounts ofrhGH expressed in modified pBAD24 vector (pBAD24M)in comparison to the original pBAD24 vector. While thelevels of rhGH reached merely 75 mg l-1 with pBAD24 in abioreactor, it reached ~1860 mg l-1 with pBAD24M vectorunder similar conditions as judged by densitometry ofproteins resolved by sodium dodecyl sulfate–polyacrylamidegel electrophoresis (SDS–PAGE). The rhGH protein wassuccessfully purified from inclusion bodies after ureadenaturation by two simple ion-exchange chromatographysteps with an overall recovery of 40% amounting to ~750mg l-1 of purified hGH which is the highest reported yield ofpurified rhGH to date. Such a purified bacterial derivedrhGH was characterized by N-terminal sequence, CDspectra studies, mass fingerprint analysis and analysis onAgilent 2100 bioanalyzer. The bioactivity of the purifiedrhGH was comparable with the commercially available hGH(somatotropin).
  Journal of Microbial & Biochemical Technology. 01/2010;
• Source

  Available from: Sriram Padmanabhan

  Article: Expression and Purification of SAK-fused Human Interferon Alpha in Escherichia coli

  Salunkhe Shardul, Prasad Bhaskarjyoti, Sabnis-Prasad Ketaki, Apte-Deshpande Anjali, Padmanabhan Sriram
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  ABSTRACT: A method for improved refolding and purification of E.coli derived human Interferon -a (rhIFN a2b) from inclusionbodies as a Staphylokinase (SAK) fusion protein isdescribed. Such a fusion protein did not require the supplementationof rare codons for expression and was found tobe stable at 37oC. The optimal conditions of refolding involvedthe use of a mild denaturating agent without theneed for any other agents to prevent aggregation. The SAKrhIFNa2b fusion protein was successfully purified usingtwo steps of purification and was cleaved using enterokinaseinto two fragments namely SAK and IFN. Both theproteins were found to be biologically active showingproper folding of both the fusion partners. The cleavedIFN showed similar retention time on RP-HPLC as thebacterial derived untagged purified IFN as well as similarmolecular weight on Agilent 2100 Bioanalyzer indicatingthe right processing of the IFN after enterokinase cleavage.The expression levels of SAK-IFN were found to betwo folds higher than that observed with untagged IFNunder similar experimental conditions.
  Journal of Microbial & Biochemical Technology. 01/2009;