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Enhanced expression of recombinant proteins utilizing a modified baculovirus expression vector

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Abstract

The baculovirus expression vector system (BEVS) has been widely used for over-expressing eukaryotic proteins due to a close resemblance in post-translational modification, processing, and transportation properties of the expressed protein, to that of the mammalian cells. In comparison to the bacterial expression system, protein yield from BEVS is relatively low, resulting in higher cost of production. To improve the existing recombinant protein expression levels, baculovirus homologous region1 (hr1) was strategically integrated into the bacmid-based transfer vectors. Luciferase reporter, human Protein Kinase B-α (PKB-A), and N-terminal-modified CYP-1A2 genes were independently cloned in non-hr1 and hr1 constructs for generating respective bacmids and baculoviruses. These recombinant baculoviruses were utilized for comparing the expresion levels at varying multiplicity of infections (MOI) and time intervals in Spodoptera frugiperda (Sf21) or Trichoplusia ni (Tni) insect cell lines. Targeted insertion of hr1 upstream to CYP-1A2, PKB-A, and Luciferase genes, compared to the non-hr1 sets, led to 3-, 3.5-, and 4.5-fold increase in the resultant protein levels, respectively. Moreover, at equal protein concentration, the corresponding activity and inhibition characteristics of these high expression hr1 sets were comparable to that of the respective non-hr1 sets. Utilization of this modified baculovirus expression construct offers significant advantage of producing recombinant proteins in a cost-effective manner for various biotechnological and therapeutic applications.

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Acknowledgments

We thankfully acknowledge Prof. Dr. S.E. Hasnain, Vice-Chancellor, University of Hyderabad, Hyderabad, India, for valuable suggestions. We thank Dr. Pradip Kumar Bhatnagar, Head, New Drug Discovery Research and our fellow scientists at Molecular Technology, Pharmacology, and Metabolism & Pharmacokinetics Departments, Ranbaxy, India, for their active involvement in this study. Ranbaxy Research Laboratories, India, is thankfully acknowledged for funding this research work.

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Authors and Affiliations

  1. Department of Molecular Technology, New Drug Discovery Research, Ranbaxy Research Laboratories-R&D-3, 20-Sector 18 Udyog Vihar, Gurgaon, 122001, India

    Prabhakar Tiwari, Shalini Saini, Sanatan Upmanyu, Kulvinder Singh Saini & Sudhir Sahdev

  2. Department of Metabolism and Pharmacokinetics, New Drug Discovery Research, Ranbaxy Research Laboratories, Gurgaon, India

    Biju Benjamin

  3. Department of Pharmacology, New Drug Discovery Research, Ranbaxy Research Laboratories, Gurgaon, India

    Ruchi Tandon

Authors
  1. Prabhakar Tiwari
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  2. Shalini Saini
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  3. Sanatan Upmanyu
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  4. Biju Benjamin
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  5. Ruchi Tandon
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  6. Kulvinder Singh Saini
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  7. Sudhir Sahdev
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Correspondence to Sudhir Sahdev.

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Prabhakar Tiwari and Shalini Saini contributed equally.

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Tiwari, P., Saini, S., Upmanyu, S. et al. Enhanced expression of recombinant proteins utilizing a modified baculovirus expression vector. Mol Biotechnol 46, 80–89 (2010). https://doi.org/10.1007/s12033-010-9284-3

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