Expression and characterization of Rv2430c, a novel immunodominant antigen of Mycobacterium tuberculosis

Protein Expression and Purification
Rakesh K ChoudharySeyed E Hasnain

Abstract

About 10% of the coding sequence of Mycobacterium tuberculosis corresponds to hitherto unknown members of the PE and PPE protein families which display significant sequence and length variation at their C-terminal region. It has been suggested that this could possibly represent a rich source of antigenic variation within the pathogen. We describe the purification and biophysical characterization of the recombinant PPE protein coded by hypothetical ORF Rv2430c, a member of the PPE gene family that was earlier shown to induce a strong B cell response. Expression of the recombinant PPE protein in Escherichia coli led to its localization in inclusion bodies and subsequent refolding using dialysis after its extraction from the same resulted in extensive precipitation. Therefore, an on-column refolding strategy was used, after which the protein was found to be in the soluble form. CD spectrum of the recombinant protein displayed predominantly alpha helical content (81%) which matched significantly with in silico and web-based secondary structure predictions. Furthermore, fluorescence emission spectra revealed that aromatic amino acids are buried inside the protein, which are exposed to aqueous environment under 8M urea. These results...Continue Reading

References

Aug 14, 2001·Infection and Immunity·G Delogu, M J Brennan
Apr 23, 2002·Molecular Microbiology·Sayera BanuStewart T Cole
May 23, 2003·Protein Science : a Publication of the Protein Society·Volety SrinivasCh Mohan Rao
Aug 5, 2003·Proceedings of the National Academy of Sciences of the United States of America·John L DahlClifton E Barry
Oct 24, 2003·Infection and Immunity·Rakesh Kumar ChoudharySeyed E Hasnain

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Citations

Apr 9, 2008·Nature Reviews. Microbiology·Niyaz AhmedSeyed E Hasnain
Dec 18, 2010·Astrobiology·Gene D McDonald, Michael C Storrie-Lombardi
Feb 15, 2011·Clinical & Developmental Immunology·Samantha L Sampson
Nov 23, 2006·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Xindu Geng, Chaozhan Wang
Oct 7, 2004·Current Opinion in Biotechnology·Alois JungbauerRobert Schlegl
Jul 20, 2007·Scandinavian Journal of Immunology·F AbebeG Bjune
Sep 26, 2019·Journal of Medical Microbiology·Ruchi RastogiJagdeep Kaur
Jun 25, 2021·Biochemical and Biophysical Research Communications·Shruti Srivastava, Sangita Mukhopadhyay

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