Abstract
A spontaneous mutant of Methanothermobacter thermautotrophicus resistant toward the ATP-synthase inhibitor N,N′-dicyclohexylcarbodiimide (DCCD) was isolated. DCCD normally inhibits methanogenic electron-transport-driven ATP synthesis, however, the DCCD-resistant strain exhibited methanogenesis in the presence of 300 μmol/L DCCD. Total ATP synthesis was shown to be higher in the mutant strain, both in the presence and absence of DCCD. These results suggested a modification in the ATP-synthesizing system of the mutant strain. Using Blue Native PAGE combined with MALDI TOF/TOF mass spectrometry, increased concentrations of both the A1 and Ao subcomplexes of the A1Ao-type synthase were identified in the mutant strain. However, no alterations were found in the structural genes (atp) for the A1Ao ATP synthase. The results imply that DCCD resistance is a consequence of increased A1Ao ATP synthase expression, and suggest that genes involved in regulating synthase expression are responsible for DCCD resistance.
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Abbreviations
- DCCD:
-
N,N′-dicyclohexylcarbodiimide
- WT:
-
wild-type (strain)
References
Azzi A., Casey R.P., Nalez M.J.: The effect of N,N′-dicyclohexylcarbodiimide on enzymes of bioenergetic relevance. Biochim.Biophys. Acta768, 209–226 (1984).
Blaut M., Gottschalk G.: Coupling of ATP synthesis and methane formation from methanol and molecular hydrogen in Methanosarcina barkeri. Eur.J.Biochem.141, 217–222 (1984).
Deppenmeyer U., Müller V., Gottschalk G.: Pathways of energy conservation in methanogenic archaea. Arch.Microbiol.165, 149–163 (1996).
Doddema H.J., Hutten T.J., Van Der Drift C., Vogels G.D.: ATP hydrolysis and synthesis by the membrane bound ATP synthase complex of Methanobacterium thermoautotrophicum. J.Bacteriol.136, 19–23 (1978).
Farhoud M.H., Wessels H.J., Steenbakkers P.J., Mattijssen S., Wevers R.A., Van Engelen B.G., Jetten M.S., Smeitinkj J.A., Van Den Heuvel L.P., Keltjens J.T.: Protein complexes in the archaeon Methanothermobacter thermautotrophicus analyzed by blue native/SDS-PAGE and mass spectrometry. Mol.Cell Proteomics4, 1653–1663 (2005).
Kaesler B., Schönheit P.: The sodium cycle in methanogenesis. CO2 reduction to the formaldehyde level in methanogenic bacteria is driven by a primary electrochemical potential of Na+ generated by formaldehyde reduction to CH4. Eur.J.Biochem.186,309–316 (1989).
Majernik A.I., Chong J.P.J.: A conserved mechanism for replication origin recognition and binding in archaea. Biochem.J.409, 511–518 (2008).
Nováková Z., Šurín S., Blaško J., Majerník A.I., Šmigáň P.: Membrane proteins and squalene-hydrosqualene profile in methanoarchaeon Methanothermobacter thermautotrophicus resistant to N,N′-dicyclohexylcarbodiimide. Folia Microbiol.53, 237–240 (2008).
Šmigáň P., Friederová A., Rusňák P., Greksák M.: Effect of 2,4-dinitrophenol and ionophores on growth and methanogenesis in Methanobacterium thermoautotrophicum. Folia Microbiol.29, 353–358 (1984).
Šmigáň P., Horovská Ľ., Greksák M.: Na+-driven ATP synthesis in Methanobacterium thermoautotrophicum can be modulated with sodium ion concentration in the growth medium. FEBS Lett.242, 85–88 (1988).
Šmigáň P., Rusnák P., Greksák M., Zhilina T.N., Zavarzin G.A.: Mode of sodium ion action on methanogenesis and ATPase of the moderate halophilic methanogenic bacterium Methanohalophilus halophilus. FEBS Lett.300, 193–196 (1992).
Šmigáň P., Majerník A., Greksák M.: Na+-driven ATP synthesis in Methanobacterium thermoautotrophicum and its differentiation from H+-driven ATP synthesis by rhodamine 6G. FEBS Lett.349, 424–428 (1994).
Šmigáň P., Majerník A., Polák P., Hapala I., Greksák M.: The presence of H+- and Na+-translocating ATPases in Methanobacterium thermoautotrophicum and their possible function under alkaline conditions. FEBS Lett.371, 119–122 (1995).
Šmigáň P., Polák P., Majerník A., Greksák M.: Isolation and characterization of a neomycin-resistant mutant of Methanobacterium thermoautotrophicum with a lesion in Na+-translocating ATPase (synthase). FEBS Lett.420, 93–96 (1997).
Smith D.R., Doucette-Stamm L.A., Deloughery C., Lee H., Dubois J., Aldredge T., Bashirzadeh R., Blakel D., Cook R., Gilbert K., Harrison D., Hoang L., Keagle P., Lumm W., Potheir B., Qiu D., Spadafora R., Vicaire R., Wang Y., Wierzbowsky J., Gibson R., Jiwani N., Caruso A., Bush D., Safer H., Patwell D., Prabhakar S., Mcdougall S., Shimer G., Goyal A., Pietrokowski S., Curchen G.M., Daniels C.J., Mao J.I., Rice P., Nolling J., Reeve J.N.: Complete genome sequence of Methanobacterium thermoautotrophicum ΔH: functional analysis and comparative genomics. J.Bacteriol.179, 7135–7155 (1997).
Šurín S., Čuboňová Ľ., Majerník A., Šmigáň P.: Amiloride resistance in the methanoarcheon Methanothermobacter thermoautotrophicus: characterization of membrane-associated proteins. Folia Microbiol.51, 313–316 (2006).
Šurín S., Čuboňová Ľ., Majerník A., Mcdermott P., Chong J.P.J., Šmigáň P.: Isolation and characterization of an amiloride-resistant mutant of Methanothermobacter thermautotrophicus possessing a defective Na+/H+-antiport. FEMS Microbiol.Lett.269, 301–308 (2007).
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Nováková, Z., Majerník, A.I., Bobáľová, J. et al. Isolation and characterization of a N,N′-dicyclohexylcarbodiimide-resistant mutant of Methanothermobacter thermautotrophicus with alterations to the ATP synthesis machinery. Folia Microbiol 54, 483–486 (2009). https://doi.org/10.1007/s12223-009-0068-8
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DOI: https://doi.org/10.1007/s12223-009-0068-8