Skip to main content
SpringerLink
Log in

Mycobacterium tuberculosis Cell Division Protein, FtsE, is an ATPase in Dimeric Form

  • Published:
The Protein Journal Aims and scope Submit manuscript

Abstract

FtsE is one of the earliest cell division proteins that assembles along with FtsX at the mid-cell site during cell division in Escherichia coli. Both these proteins are highly conserved across diverse bacterial genera and are predicted to constitute an ABC transporter type complex, in which FtsE is predicted to bind ATP and hydrolyse it, and FtsX is predicted to be an integral membrane protein. We had earlier reported that the MtFtsE of the human pathogen, Mycobacterium tuberculosis, binds ATP and interacts with MtFtsX on the cell membrane of M. tuberculosis and E. coli. In this study, we demonstrate that MtFtsE is an ATPase, the active form of which is a dimer, wherein the participating monomers are held together by non-covalent interactions, with the Cys84 of each monomer present at the dimer interface. Under oxidising environment, the dimer gets stabilised by the formation of Cys84–Cys84 disulphide bond. While the recombinant MtFtsE forms a dimer on the membrane of E. coli, the native MtFtsE seems to be in a different conformation in the M. tuberculosis membrane. Although disulphide bridges were not observed on the cytoplasmic side (reducing environment) of the membrane, the two participating monomers could be isolated as dimers held together by non-covalent interactions. Taken together, these findings show that MtFtsE is an ATPase in the non-covalent dimer form, with the Cys84 of each monomer present in the reduced form at the dimer interface, without participating in the dimerisation or the catalytic activity of the protein.

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
Fig. 6
Fig. 7

Similar content being viewed by others

Abbreviations

ABC:

ATP binding cassette

ATP:

Adenosine triphosphate

ADP:

Adenosine diphosphate

Fts:

Filamenting temperature sensitive

NBD:

Nucleotide binding domain

TMD:

Trans membrane domain

DTT:

Dithiothreitol

IPTG:

Isopropylthio-β-d-galactoside

LB broth:

Luria–Bertani broth

PBS:

Phosphate buffered saline

PMSF:

Phenyl methyl sulphonyl fluoride

CuP:

Cupric phenanthroline

DTSP:

3,3′-Dithio-bispropionic acid di (N-hydroxysuccinimide ester)

SDS-PAGE:

Sodium dodecyl sulphate-polyacrylamide gel electrophoresis

β-ΜΕ:

β-Mercaptoethanol

MBP:

Maltose binding protein

References

  1. Schmidt KL, Peterson ND, Kustusch RJ, Wissel MC, Graham B, Phillips GJ, Weiss DS (2004) A predicted ABC transporter, FtsEX, is needed for cell division in Escherichia coli. J Bacteriol 186:785–793

    Article  CAS  Google Scholar 

  2. Braibant M, Gilot P, Content J (2000) The ATP binding cassette (ABC) transport systems of Mycobacterium tuberculosis. FEMS Microbiol Rev 24:449–467

    Article  CAS  Google Scholar 

  3. Ambudkar SV, Kim IW, Xia D, Sauna ZE (2006) The A-loop, a novel conserved aromatic acid subdomain upstream of the Walker A motif in ABC transporters, is critical for ATP binding. FEBS Lett 580:1049–1055

    Article  CAS  Google Scholar 

  4. Smith PC, Karpowich N, Millen L, Moody JE, Rosen J, Thomas PJ, Hunt JF (2002) ATP binding to the motor domain from an ABC transporter drives formation of a nucleotide sandwich dimer. Mol Cell 10:139–149

    Article  CAS  Google Scholar 

  5. Loo TW, Bartlett MC, Clarke DM (2002) The “LSGGQ” motif in each nucleotide-binding domain of human P-glycoprotein is adjacent to the opposing walker A sequence. J Biol Chem 277:41303–41306

    Article  CAS  Google Scholar 

  6. de Leeuw E, Graham B, Phillips GJ, ten Hagen-Jongman CM, Oudega B, Luirink J (1999) Molecular characterisation of Escherichia coli FtsE and FtsX. Mol Microbiol 31:983–993

    Article  Google Scholar 

  7. Mir MA, Rajeswari HS, Veeraraghavan U, Ajitkumar P (2006) Molecular characterisation of ABC transporter type FtsE and FtsX proteins of Mycobacterium tuberculosis. Arch Microbiol 185:147–158

    Article  CAS  Google Scholar 

  8. Sham LT, Barendt SM, Kopecky KE, Winkler ME (2011) Essential PcsB putative peptidoglycan hydrolase interacts with the essential FtsXSpn cell division protein in Streptococcus pneumoniae D39. Proc Natl Acad Sci USA 108:E1061–E1069

    Article  Google Scholar 

  9. Yang DC, Peters NT, Parzych KR, Uehara T, Markovski M, Bernhardt TG (2011) An ATP-binding cassette transporter-like complex governs cell-wall hydrolysis at the bacterial cytokinetic ring. Proc Natl Acad Sci USA 108:E1052–E1060

    Article  Google Scholar 

  10. Meisner J, Montero Llopis P, Sham LT, Garner E, Bernhardt TG, Rudner DZ (2013) FtsEX is required for CwlO peptidoglycan hydrolase activity during cell wall elongation in Bacillus subtilis. Mol Microbiol 89:1069–1083

    Article  CAS  Google Scholar 

  11. Arends SJ, Kustusch RJ, Weiss DS (2009) ATP-binding site lesions in FtsE impair cell division. J Bacteriol 191:3772–3784

    Article  CAS  Google Scholar 

  12. Gill DR, Salmond GP (1987) The Escherichia coli cell division proteins FtsY, FtsE and FtsX are inner membrane-associated. Mol Gen Genet 210:504–508

    Article  CAS  Google Scholar 

  13. Ukai H, Matsuzawa H, Ito K, Yamada M, Nishimura A (1998) ftsE(Ts) affects translocation of K+-pump proteins into the cytoplasmic membrane of Escherichia coli. J Bacteriol 180:3663–3670

    CAS  Google Scholar 

  14. Reddy M (2007) Role of FtsEX in cell division of Escherichia coli: viability of ftsEX mutants is dependent on functional SufI or high osmotic strength. J Bacteriol 189:98–108

    Article  CAS  Google Scholar 

  15. Guzman LM, Belin D, Carson MJ, Beckwith J (1995) Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol 177:4121–4130

    CAS  Google Scholar 

  16. Randerath E, Randerath K (1967) Ion-exchange thin-layer chromatography: XVI. Techniques for preparation and analysis of oligonucleotides. J Chromatogr 31:485–499

    Article  CAS  Google Scholar 

  17. Oudot C, Jaquinod M, Cortay JC, Cozzone AJ, Jault JM (1999) The isocitrate dehydrogenase kinase/phosphatase from Escherichia coli is highly sensitive to in vitro oxidative conditions role of cysteine67 and cysteine108 in the formation of a disulfide-bonded homodimer. Eur J Biochem 262:224–229

    Article  CAS  Google Scholar 

  18. Zhang Y, Fillingame RH (1995) Subunits coupling H+ transport and ATP synthesis in the Escherichia coli ATP synthase. Cys–Cys cross-linking of F1 subunit epsilon to the polar loop of F0 subunit c. J Biol Chem 270:24609–24614

    Article  CAS  Google Scholar 

  19. Hunke S, Mourez M, Jehanno M, Dassa E, Schneider E (2000) ATP modulates subunit-subunit interactions in an ATP-binding cassette transporter (MalFGK2) determined by site-directed chemical cross-linking. J Biol Chem 275:15526–15534

    Article  CAS  Google Scholar 

  20. Kobashi K (1968) Catalytic oxidation of sulfhydryl groups by o-phenanthroline copper complex. Biochim Biophys Acta 158:239–245

    Article  CAS  Google Scholar 

  21. Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN, Weissig H, Shindyalov IN, Bourne PE (2000) The protein data bank. Nucleic Acids Res 28:235–242

    Article  CAS  Google Scholar 

  22. Diederichs K, Diez J, Greller G, Muller C, Breed J, Schnell C, Vonrhein C, Boos W, Welte W (2000) Crystal structure of MalK, the ATPase subunit of the trehalose/maltose ABC transporter of the archaeon Thermococcus litoralis. EMBO J 19:5951–5961

    Article  CAS  Google Scholar 

  23. Laskowski RA, Rullmannn JA, MacArthur MW, Kaptein R, Thornton JM (1996) AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR. J Biomol NMR 8:477–486

    Article  CAS  Google Scholar 

  24. Cohen GH (1997) ALIGN: a program to superimpose protein coordinates, accounting for insertions and deletions. J Appl Cryst 30:1160–1161

    Article  CAS  Google Scholar 

  25. Hubbard SJ, Campbell SF, Thornton JM (1991) Molecular recognition: conformational analysis of limited proteolytic sites and serine proteinase protein inhibitors. J Mol Biol 220:507–530

    Article  CAS  Google Scholar 

  26. Shi J, Blundell TL, Mizuguchi K (2001) FUGUE: sequence-structure homology recognition using environment-specific substitution tables and structure-dependent gap penalties. J Mol Biol 310:243–257

    Article  CAS  Google Scholar 

  27. Hopfner KP, Karcher A, Shin DS, Craig L, Arthur LM, Carney JP, Tainer JA (2000) Structural biology of Rad50 ATPase: ATP-driven conformational control in DNA double-strand break repair and the ABC-ATPase superfamily. Cell 101:789–800

    Article  CAS  Google Scholar 

  28. Chen J, Lu G, Lin J, Davidson AL, Quiocho FA (2003) A tweezers-like motion of the ATP-binding cassette dimer in an ABC transport cycle. Mol Cell 12:651–661

    Article  CAS  Google Scholar 

  29. Collet JF, Bardwell JC (2002) Oxidative protein folding in bacteria. Mol Microbiol 44:1–8

    Article  CAS  Google Scholar 

  30. Nikaido K, Liu PQ, Ames GF (1997) Purification and characterization of HisP, the ATP-binding subunit of a traffic ATPase (ABC transporter), the histidine permease of Salmonella typhimurium. Solubility, dimerisation, and ATPase activity. J Biol Chem 272:27745–27752

    Article  CAS  Google Scholar 

  31. Schneider E, Hunke S (1998) ATP-binding-cassette (ABC) transport systems: functional and structural aspects of the ATP-hydrolysing subunits/domains. FEMS Microbiol Rev 22:1–20

    Article  CAS  Google Scholar 

  32. Reich-Slotky R, Panagiotidis C, Reyes M, Shuman HA (2000) The detergent-soluble maltose transporter is activated by maltose binding protein and verapamil. J Bacteriol 182:993–1000

    Article  CAS  Google Scholar 

  33. Liu PQ, Ames GF (1998) In vitro disassembly and reassembly of an ABC transporter, the histidine permease. Proc Natl Acad Sci USA 95:3495–3500

    Article  CAS  Google Scholar 

  34. Heras B, Shouldice SR, Totsika M, Scanlon MJ, Schembri MA, Martin JL (2009) DSB proteins and bacterial pathogenicity. Nat Rev Microbiol 7:215–225

    Article  CAS  Google Scholar 

  35. McIntosh PR, Freedman RB (1980) Characteristics of a copper-dependent cross-linking reaction between two forms of cytochrome P-450 in rabbit-liver microsomal membranes. Biochem J 187:227–237

    CAS  Google Scholar 

  36. Tsumoto K, Umetsu M, Kumagai I, Ejima D, Philo JS, Arakawa T (2004) Role of arginine in protein refolding, solubilisation, and purification. Biotechnol Prog 20:1301–1308

    Article  CAS  Google Scholar 

  37. Nikaido K, Ames GF (1999) One intact ATP-binding subunit is sufficient to support ATP hydrolysis and translocation in an ABC transporter, the histidine permease. J Biol Chem 274:26727–26735

    Article  CAS  Google Scholar 

  38. Chen CA, Cowan JA (2003) Characterization of the soluble domain of the ABC7 type transporter Atm1. J Biol Chem 278:52681–52688

    Article  CAS  Google Scholar 

  39. Morbach S, Tebbe S, Schneider E (1993) The ATP-binding cassette (ABC) transporter for maltose/maltodextrins of Salmonella typhimurium. Characterisation of the ATPase activity associated with the purified MalK subunit. J Biol Chem 268:18617–18621

    CAS  Google Scholar 

  40. Treptow NA, Shuman HA (1985) Genetic evidence for substrate and periplasmic-binding-protein recognition by the MalF and MalG proteins, cytoplasmic membrane components of the Escherichia coli maltose transport system. J Bacteriol 163:654–660

    CAS  Google Scholar 

  41. Bishop L, Agbayani R Jr, Ambudkar SV, Maloney PC, Ames GF (1989) Reconstitution of a bacterial periplasmic permease in proteoliposomes and demonstration of ATP hydrolysis concomitant with transport. Proc Natl Acad Sci USA 86:6953–6957

    Article  CAS  Google Scholar 

  42. Anderson MP, Berger HA, Rich DP, Gregory RJ, Smith AE, Welsh MJ (1991) Nucleoside triphosphates are required to open the CFTR chloride channel. Cell 67:775–784

    Article  CAS  Google Scholar 

  43. Liu CE, Ames GF (1997) Characterisation of transport through the periplasmic histidine permease using proteoliposomes reconstituted by dialysis. J Biol Chem 272:859–866

    Article  CAS  Google Scholar 

  44. Hobson AC, Weatherwax R, Ames GF (1984) ATP-binding sites in the membrane components of histidine permease, a periplasmic transport system. Proc Natl Acad Sci USA 81:7333–7337

    Article  CAS  Google Scholar 

  45. Higgins CF, Hiles ID, Whalley K, Jamieson DJ (1985) Nucleotide binding by membrane components of bacterial periplasmic binding protein-dependent transport systems. EMBO J 4:1033–1039

    CAS  Google Scholar 

  46. Arumugam M, Ajitkumar P (2012) Histidine 117 in the His-Gly-Ser-Asp motif is required for the biochemical activities of nucleoside diphosphate kinase of Mycobacterium smegmatis. Open Biochem J 6:71–77

    Article  CAS  Google Scholar 

Download references

Acknowledgments

MAM thanks Prof. Keith M. Derbyshire, Division of Genetics, Wadsworth Center, Albany, New York, USA, for critical comments on the manuscript. This work was carried out using the ICMR research Grant No. 63/72/2001-BMS to PA and the infrastructural facilities provided by the DBT-supported Genomics Initiative on Microbial Pathogens–Structural Genomics Initiative in the Division of Biological Sciences, Indian Institute of Science, the DST-FIST and UGC-CAS at the Department of Microbiology and Cell Biology, Indian Institute of Science. The authors acknowledge the facilities at the erstwhile DBT-funded Bioinformatics Centre at IISc.

Author information

Authors and Affiliations

  1. Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, 560012, India

    Mushtaq Ahmad Mir, Muthu Arumugam, Haryadi S. Rajeswari & Parthasarathi Ajitkumar

  2. Department of Physics and Bioinformatics Centre, Indian Institute of Science, Bangalore, 560012, India

    Sukanta Mondal & Suryanarayanarao Ramakumar

  3. Division of Genetics, New York State Department of Health, Wadsworth Center, Albany, NY, 12201, USA

    Mushtaq Ahmad Mir

  4. CSIR-National Institute for Interdisciplinary Science and Technology, Thiruvananthapuram, 695019, Kerala, India

    Muthu Arumugam

  5. Biological Sciences, Birla Institute of Technology and Science, Pilani K. K. Birla Goa Campus, Zuarinagar, Goa, 403726, India

    Sukanta Mondal

Authors
  1. Mushtaq Ahmad Mir
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Muthu Arumugam
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sukanta Mondal
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Haryadi S. Rajeswari
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Suryanarayanarao Ramakumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Parthasarathi Ajitkumar
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Parthasarathi Ajitkumar.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Mir, M.A., Arumugam, M., Mondal, S. et al. Mycobacterium tuberculosis Cell Division Protein, FtsE, is an ATPase in Dimeric Form. Protein J 34, 35–47 (2015). https://doi.org/10.1007/s10930-014-9593-7

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10930-014-9593-7

Keywords

Search

Navigation