Skip to main content

Advertisement

SpringerLink
Log in

Prokaryotic Expression, Identification and Bioinformatics Analysis of the Mycobacterium tuberculosis Rv3807c Gene Encoding the Putative Enzyme Committed to Decaprenylphosphoryl-d-arabinose Synthesis

  • Original Article
  • Published:
Indian Journal of Microbiology Aims and scope Submit manuscript

Abstract

Decaprenylphosphoryl-d-arabinofuranosyl (DPA), the immediate donor for the polymerized d-Araf residues of mycobacterial arabinan, is synthesized from 5-phosphoribose-1-diphosphate (PRPP) in three-step reactions. (i) PRPP is transferred to decaprenyl-phosphate (DP) to form decaprenylphosphoryl-d-5-phosphoribose (DPPR). (ii) DPPR is dephosphorylated to form decaprenylphosphoryl-d-ribose (DPR). (iii) DPR is formed to DPA by the epimerase. Mycobacterium tuberculosis Rv3806c and heteromeric Rv3790/Rv3791 have been identified as the PRPP: decaprenyl-phosphate 5-phosphoribosyltransferase and the epimerase respectively. Rv3807c, however, as the candidate of phospholipid phosphatase, catalyzing the biosynthesis of decapreny-l-phosphoryl-ribose (DPR) from decaprenylphosphoryl-β-d-5-phosphoribose by dephosphorylating, has no direct experimental evidence of its essentiality in any species of mycobacterium. In this study, Rv3807c gene was amplified from the genome of M. tuberculosis H37Rv by PCR, and was successfully expressed in Escherichia coli BL21 (DE3) via the recombinant plasmid pColdII-Rv3807c. The resulting protein with the 6× His-tag was identified by SDS-PAGE and Western blotting. The protein was predicted through bioinformatics to contain three transmembrane domains, the N-terminal peptide, and a core structure with phosphatidic acid phosphatase type2/haloperoxidase. This study provides biochemical and bioinformatics evidence for the importance of Rv3807c in mycobacteria, and further functional studies will be conducted for validating Rv3807c as a promising phospholipid phosphatase in the synthetic pathway of DPA.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

BCIP:

5-Bromo-4-chloro-3-indolylphosphate

DP:

Decaprenyl-phosphate

DPA:

Decaprenyl-phospho-d-arabinose

DPPR:

Decaprenylphosphoryl-d-5-phosphoribose

DPR:

Decaprenylphosphoryl-d-ribose

NBT:

Nitroblue tetrazolium

Ni-NTA:

Ni-nitrilotriacetic acid

EDTA:

Ethylenediaminetetraacetic acid

PAGE:

Polyacrylamide gel electrophoresis

PRPP:

5-Phosphoribose-1-diphosphate

SDS:

Sodium dodecyl sulfate

References

  1. Brennan PJ (2003) Structure, function, and biogenesis of the cell wall of Mycobacterium tuberculosis. Tuberculosis (Edinb) 83(1–3):91–97

    Article  CAS  Google Scholar 

  2. Crick DC, Schulbach MC, Zink EE, Macchia M, Barontini S, Besra GS et al (2000) Polyprenyl phosphate biosynthesis in Mycobacterium tuberculosis and Mycobacterium smegmatis. J Bacteriol 182(20):5771–5778

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  3. Crick DC, Mahapatra S, Brennan PJ (2001) Biosynthesis of the arabinogalactan–peptidoglycan complex of Mycobacterium tuberculosis. Glycobiology 11(9):107R–118R

    Article  CAS  PubMed  Google Scholar 

  4. Scherman M, Weston A, Duncan K, Whittington A, Upton R, Deng L et al (1995) Biosynthetic origin of mycobacterial cell wall arabinosyl residues. J Bacteriol 177(24):7125–7130

    CAS  PubMed Central  PubMed  Google Scholar 

  5. Wolucka BA, McNeil MR, de Hoffmann E, Chojnacki T, Brennan PJ (1994) Recognition of the lipid intermediate for arabinogalactan/arabinomannan biosynthesis and its relation to the mode of action of ethambutol on mycobacteria. J Biol Chem 269(37):23328–23335

    CAS  PubMed  Google Scholar 

  6. Crellin PK, Brammananth R, Coppel RL (2011) Decaprenylphosphoryl-beta-d-ribose 2′-epimerase, the target of benzothiazinones and dinitrobenzamides, is an essential enzyme in Mycobacterium smegmatis. PLoS One 6(2):e16869. doi:10.1371/journal.pone.0016869

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Horita Y, Takii T, Kuroishi R, Chiba T, Ogawa K, Kremer L et al (2011) Synthesis and evaluation of anti-tubercular activity of new dithiocarbamate sugar derivatives. Bioorg Med Chem Lett 21(3):899–903. doi:10.1016/j.bmcl.2010.12.084

    Article  CAS  PubMed  Google Scholar 

  8. Buroni S, Pasca MR, de Jesus Lopes Ribeiro AL, Degiacomi G, Molteni E, Riccardi G (2012) Antituberculars which target decaprenylphosphoryl-β-d-ribofuranose 2′-oxidase DprE1: state of art. Appl Microbiol Biotechnol 94(4):907–916. doi:10.1007/s00253-012-4013-4

    Article  CAS  PubMed  Google Scholar 

  9. Huang H, Scherman MS, D’Haeze W, Vereecke D, Holsters M, Crick DC et al (2005) Identification and active expression of the Mycobacterium tuberculosis gene encoding 5-phospho-α-d-ribose-1-diphosphate: decaprenyl-phosphate 5-phosphoribosyltransferase, the first enzyme committed to decaprenylphosphoryl-d-arabinose synthesis. J Biol Chem 280(26):24539–24543. doi:10.1074/jbc.M504068200

    Article  CAS  PubMed  Google Scholar 

  10. Jiang T, He L, Zhan Y, Zang S, Ma Y, Zhao X et al (2011) The effect of MSMEG_6402 gene disruption on the cell wall structure of Mycobacterium smegmatis. Microb Pathog 51(3):156–160. doi:10.1016/j.micpath.2011.04.005

    Article  CAS  PubMed  Google Scholar 

  11. Wolucka BA (2008) Biosynthesis of d-arabinose in mycobacteria—a novel bacterial pathway with implications for antimycobacterial therapy. FEBS J 275(11):2691–2711. doi:10.1111/j.1742-4658.2008.06395.x

    Article  CAS  PubMed  Google Scholar 

  12. Mikusova K, Huang H, Yagi T, Holsters M, Vereecke D, D’Haeze W et al (2005) Decaprenylphosphoryl arabinofuranose, the donor of the d-arabinofuranosyl residues of mycobacterial arabinan, is formed via a two-step epimerization of decaprenylphosphoryl ribose. J Bacteriol 187(23):8020–8025. doi:10.1128/JB.187.23.8020-8025.2005

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  13. Eoh H, Brown AC, Buetow L, Hunter WN, Parish T, Kaur D et al (2007) Characterization of the Mycobacterium tuberculosis 4-diphosphocytidyl-2-C-methyl-d-erythritol synthase: potential for drug development. J Bacteriol 189(24):8922–8927. doi:10.1128/JB.00925-07

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  14. Sassetti CM, Boyd DH, Rubin EJ (2003) Genes required for mycobacterial growth defined by high density mutagenesis. Mol Microbiol 48(1):77–84

    Article  CAS  PubMed  Google Scholar 

  15. Slayden RA, Jackson M, Zucker J, Ramirez MV, Dawson CC, Crew R et al (2013) Updating and curating metabolic pathways of TB. Tuberculosis (Edinb) 93(1):47–59. doi:10.1016/j.tube.2012.11.001

    Article  CAS  Google Scholar 

  16. Carman GM, Han GS (2006) Roles of phosphatidate phosphatase enzymes in lipid metabolism. Trends Biochem Sci 31(12):694–699. doi:10.1016/j.tibs.2006.10.003

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  17. Dong X, Bhamidi S, Scherman M, Xin Y, McNeil MR (2006) Development of a quantitative assay for mycobacterial endogenous arabinase and ensuing studies of arabinase levels and arabinan metabolism in Mycobacterium smegmatis. Appl Environ Microbiol 72(4):2601–2605. doi:10.1128/AEM.72.4.2601-2605.2006

    Article  CAS  PubMed Central  PubMed  Google Scholar 

Download references

Acknowledgments

This study is supported by the grants from National Natural Science Foundation of China (30970647).

Conflict of interest

The authors declare that they have no conflicts of interest.

Author information

Authors and Affiliations

  1. Department of Biotechnology, Dalian Medical University, Dalian, 116044, People’s Republic of China

    Lina Cai, Tao Jiang, Juanjuan Qiu, Lawrence Owusu & Yi Xin

  2. TB Laboratory of Shenyang Chest Hospital, Shenyang, 110044, People’s Republic of China

    Xiaojiao Zhao

  3. Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian, 116044, People’s Republic of China

    Yufang Ma

  4. Department of Pathology, Dalian Medical University, Dalian, 116044, People’s Republic of China

    Bo Wang

Authors
  1. Lina Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xiaojiao Zhao
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tao Jiang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Juanjuan Qiu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Lawrence Owusu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yufang Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Bo Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Yi Xin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Yi Xin.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cai, L., Zhao, X., Jiang, T. et al. Prokaryotic Expression, Identification and Bioinformatics Analysis of the Mycobacterium tuberculosis Rv3807c Gene Encoding the Putative Enzyme Committed to Decaprenylphosphoryl-d-arabinose Synthesis. Indian J Microbiol 54, 46–51 (2014). https://doi.org/10.1007/s12088-013-0418-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12088-013-0418-8

Keywords

Search

Navigation