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Industrial Scale Production of Recombinant Human Insulin using Escherichia coli BL-21

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Iranian Journal of Science and Technology, Transactions A: Science Aims and scope Submit manuscript

Abstract

Insulin is a well-characterized peptide produced by recombinant DNA technology that is widely used in maintaining blood glucose level in the diabetic patients. In this study, we have used vector pET-9a to express the human insulin in E. coli BL-21 DE3. The recombinant E. coli cells were cultured by fed-batch fermentation (20.0 L) process which resulted in a dry cell mass of 20 g/L. Pro-Insulin was expressed as inclusion bodies (IBs) and it was 20% of the total protein measured by densitometry. The IBs were then solubilized and refolded to get the native biological structure. The proinsulin form has been converted into active insulin by enzymatic digestion using trypsin and carboxypeptidase-B. The active insulin was purified using ion-exchange and reverse phase HPLC. The final purified insulin has achieved a purity of 98% with Des-Insulin impurity ≤ 0.5%, when analyzed in analytical HPLC as per the United States Pharmacopiea (USP) method. This optimized process of recombinant human insulin may be used as a model for insulin analogues such as glargine, aspart, lispro etc.

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Abbreviations

HPLC:

High Performance Liquid Chromatography

IBs:

Inclusion Bodies

E. coli :

Escherichia coli

USP:

United States Pharmacopeia

DNA:

Deoxyribonucleic acid

WHO:

World health organization

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Acknowledgements

The authors are grateful to Vcare Biolabs Pvt. Ltd., Hyderabad for providing facilities to carry out the work. Mr. K. Satish Babu is thankful to Management, VFSTR, India for providing opportunity to pursue his Ph.D.

Funding

No financial assistance was received either from government or any private agency for the present work.

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Authors and Affiliations

  1. Department of Biotechnology, Vignan’s Foundation for Science, Technology and Research, Vadlamudi, Guntur, 522213, Andhra Pradesh, India

    Satish Babu Kaki, Anjani Devi Chintagunta, Vijaya Ramu Dirisala & N. S. Sampath Kumar

  2. Wockhardt Research Center, Aurangabad, 431210, Maharashtra, India

    A. Naga Prasad

  3. Vcare Biolabs Pvt. Ltd., Hyderabad, Telangana, 500090, India

    S. J. K. Naidu & B. Ramesh

Authors
  1. Satish Babu Kaki
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  2. A. Naga Prasad
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  3. Anjani Devi Chintagunta
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  4. Vijaya Ramu Dirisala
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  5. N. S. Sampath Kumar
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  6. S. J. K. Naidu
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  7. B. Ramesh
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Contributions

KSB: Investigation; NPA: Conceptualization and curation; ADC: Writing-original draft; VRD: Reviewing the manuscript; NSSK: Writing-original draft and Reviewing the manuscript; SJKN: Methodology; RB: Resource.

Corresponding author

Correspondence to N. S. Sampath Kumar.

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Conflict of interest

The authors declare that they have no conflicts of interest with respect to the work described in this manuscript.

Human and animal rights statement

This article does not contain any studies with human or animal subjects.

Availability of data and material

The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions.

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Cite this article

Kaki, S.B., Naga Prasad, A., Chintagunta, A.D. et al. Industrial Scale Production of Recombinant Human Insulin using Escherichia coli BL-21. Iran J Sci Technol Trans Sci 46, 373–383 (2022). https://doi.org/10.1007/s40995-022-01269-7

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  • DOI: https://doi.org/10.1007/s40995-022-01269-7

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