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Molecular Biotechnology

, Volume 61, Issue 12, pp 905–915 | Cite as

High Yield Expression of Recombinant CD151 in E. coli and a Structural Insight into Cholesterol Binding Domain

  • Gayathri Purushothaman
  • Vijay ThiruvenkatamEmail author
Original paper

Abstract

CD151 is an abundantly expressed eukaryotic transmembrane protein on the cell surface. It is involved in cell adhesion, angiogenesis and signal transduction as well in disease conditions such as cancer and viral infections. However, the molecular mechanism of CD151 activation is poorly understood due to the lack of structural information. By considering the difficulties in expressing the membrane protein in E. coli, herein we introduce the strategic design for the effective expression of recombinant CD151 protein in E. coli with high yield, that would aid for the structural studies. CD151 having four transmembrane domain (TMD’s) along with small and a large extracellular loop (LEL) is constructed in parts to enhance the soluble expression of the protein attached with fusion tag. This has led to the high yield of the recombinant CD151 protein in the designed constructs. The recombinant CD151 protein is characterized and confirmed by western blot, CD and Mass peptide fingerprint. The molecular dynamics simulations (MDS) for the full-length CD151 shows conformational changes in the LEL of the protein in the presence and absence of cholesterol and indicate the certainty of closed and open conformation of CD151 based on cholesterol binding. The MDS results have led to the understanding of the possible underlying mechanism for the activation of the CD151 protein.

Keywords

Transmembrane protein CD151 Construct design Recombinant expression Molecular dynamics simulation Cholesterol binding domain 

Notes

Acknowledgements

The authors thank SERB Project No.: EMR/2016/001,022 for funding and IIT Gandhinagar for infrastructure. We also thank Dr Sivapriya Kirubakaran for her constant support and helpful discussions, and Althaf Shaik and Deekshi Angira for proofreading the manuscript.

Author Contributions

VT conceptualized the idea and designed the project. VT and GP designed all the experiments and protocols. GP performed all the experiments. VT and GP wrote the manuscript together.

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflicts of interest with the contents of this article.

Supplementary material

12033_2019_212_MOESM1_ESM.docx (6.4 mb)
Supplementary material 1 (DOCX 6528 kb)

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© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Discipline of Biological EngineeringIndian Institute of Technology GandhinagarGandhinagarIndia

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