Skip to main content
SpringerLink
Log in

Electrogenesis in Plasma Membrane Fraction of Halotolerant Microalga Dunaliella maritima and Effects of N,N′-Dicyclohexylcarbodiimide

  • Published:
Biochemistry (Moscow) Aims and scope Submit manuscript

Abstract

The effects of N,N′-dicyclohexylcarbodiimide (DCCD), non-specific inhibitor of various transport systems functioning in biological membranes, on Na+-transporting P-type ATPase of the green halotolerant microalga Dunaliella maritima were studied in the experiments with vesicular plasma membranes isolated from the alga cells. The effects of DCCD on electrogenic/ion transport function of the enzyme and its ATP hydrolase activity were investigated. Electrogenic/ion transport function of the enzyme was recorded as a Na+-dependent generation of electric potential on the vesicle membranes with the help of the potential-sensitive probe oxonol VI. It was found that unlike many other ion-transporting ATPases, the Na+-ATPase of D. maritima is insensitive to DCCD. This agent did not inhibit either ATP hydrolysis catalyzed by this enzyme or its transport activity. At the same time DCCD affected the ability of the vesicle membranes to maintain electric potential generated by the D. maritima Na+-ATPase. The observed effects can be explained based on the assumption that DCCD interacts with the Na+/H+ antiporter in the plasma membrane of D. maritima.

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.

Similar content being viewed by others

Abbreviations

Δψ:

transmembrane electric potential

CCCP:

carbonyl cyanide m-chlorophenylhydrazone

DCCD:

N,N′-dicyclohexylcarbodiimide

ETH157:

sodium ionophore II (N,N′-dibenzyl-N,N′-diphenyl-1,2-phenylenedioxydiacetamide)

PM:

plasma membrane

REFERENCES

  1. Boldyrev, A. A. (2001) Na/K-ATPase as an oligomeric ensemble, Biochemistry (Moscow), 66, 821-831, doi: https://doi.org/10.1023/a:1011964832767.

    Article  CAS  Google Scholar 

  2. Scheiner-Bobis, G. (2002) The sodium pump, Eur. J. Biochem., 269, 2424-2433, doi: https://doi.org/10.1046/j.1432-1033.2002.02909.x.

    Article  CAS  PubMed  Google Scholar 

  3. Gaxiola, R. A., Palmgren, M. G., and Schumacher, K. (2007) Plant proton pumps, FEBS Lett., 581, 2204-2214, doi: https://doi.org/10.1016/j.febslet.2007.03.050.

    Article  CAS  PubMed  Google Scholar 

  4. Balnokin, Y. V., and Popova, L. G. (1994) The ATP-driven Na+-pump in the plasma membrane of the marine unicellular alga Platymonas viridis, FEBS Lett., 343, 61-64, doi: https://doi.org/10.1016/0014-5793(94)80607-1.

    Article  CAS  PubMed  Google Scholar 

  5. Popova, L. G., Shumkova, G. A., Andreev, I. M., and Balnokin, Y. V. (2005) Functional identification of electrogenic Na+-translocating ATPase in the plasma membrane of the halotolerant microalga Dunaliella maritima, FEBS Lett., 579, 5002-5006, doi: https://doi.org/10.1016/j.febslet.2005.07.087.

    Article  CAS  PubMed  Google Scholar 

  6. Axelsen, K., and Palmgren, M. G. (1998) Evolution and substrate specificities in the P-type ATPase superfamily, J. Mol. Evol., 46, 84-101, doi: https://doi.org/10.1007/pl00006286.

    Article  CAS  PubMed  Google Scholar 

  7. Palmgren, M. G., and Nissen, P. (2011) P-type ATPases, Annu. Rev. Biophys., 40, 243-266, doi: https://doi.org/10.1146/annurev.biophys.093008.131331.

    Article  CAS  PubMed  Google Scholar 

  8. Pagis, L. Y., Popova, L. G., Andreev, I. M., and Balnokin, Y. V. (2003) Comparative characterization of the two primary pumps, H+-ATPase and Na+-ATPase, in the plasma membrane of the marine alga Tetraselmis viridis, Physiol. Plant., 118, 514-522, doi: https://doi.org/10.1034/j.1399-3054.2003.00113.x.

    Article  CAS  Google Scholar 

  9. Pedersen, Ch. N. S., Axelsen, K. B., Harper, J. F., and Palmgren, M. G. (2012) Evolution of plant P-type ATPases, Front. Plant Sci., 3, doi: https://doi.org/10.3389/fpls.2012.00031.

    Article  Google Scholar 

  10. Oren, A. (2005) A hundred years of Dunaliella research: 1905-2005, Saline Systems, 1, 1-14, doi: https://doi.org/10.1186/1746-1448-1-2.

    Article  Google Scholar 

  11. Gouaux, E., and MacKinnon, R. (2005) Principles of selective ion transport in channels and pumps, Science, 310, 1461-1465, doi: https://doi.org/10.1126/science.1113666.

    Article  CAS  PubMed  Google Scholar 

  12. Meier, T., Krah, A., Bond, P. J., Pogoyelov, D., Diederichs, K., and Faraldo-Gomez, J. D. (2009) Complete ion-coordination structure in the rotor ring of Na+-dependent F-ATP synthases, J. Mol. Biol., 391, 498-507, doi: https://doi.org/10.1016/j.jmb.2009.05.082.

    Article  CAS  PubMed  Google Scholar 

  13. Imagawa, T., Yamamoto, T., Kaya, Sh., Sakaguchi, K., and Taniguchi, K. (2005) Thr-774 (transmembrane segment M5), Val-920 (M8), and Glu-923 (M9) are involved in Na+-transport, and Gln-923 (M8) is essential for Na+,K+-ATPase activity, J. Biol. Chem., 280, 18736-18744, doi: https://doi.org/10.1074/jbc.M500137200.

    Article  CAS  PubMed  Google Scholar 

  14. Kaim, G., and Dimroth, P. (1994) Construction, expression and characterization of a plasmid-encoded Na+-specific ATPase hybrid consisting of Propionigenium modestum Fo-ATPase and Escherichia coli F1-ATPase, Eur. J. Biochem., 222, 615-623, doi: https://doi.org/10.1111/j.1432-1033.1994.tb18904.x.

    Article  CAS  PubMed  Google Scholar 

  15. Toei, M., and Noji, H. (2013) Single-molecule analysis of F0F1-ATP synthase inhibited by N,N′-dicyclohexylcarbodiimide, J. Biol. Chem., 288, 25717-25726, doi: https://doi.org/10.1074/jbc.M113.482455.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Ferguson, S., Keis, S., and Cook, G. M. (2006) Biochemical and molecular characterization of a Na+-translocating F1F0-ATPase from the thermoalkaliphilic bacterium Clostridium paradoxum, J. Bacteriol., 188, 5045-5054, doi: https://doi.org/10.1128/JB.00128-06.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Soontharapirakkul, K., and Incharoensakdi, A. (2010) Na+-stimulated ATPase of alkaliphilic halotolerant cyanobacterium Aphanothece halophytica translocates Na+ into proteoliposomes via Na+ uniport mechamism, BMC Biochemistry, 11, doi: https://doi.org/10.1186/1471-2091-11-30.

    Article  Google Scholar 

  18. Murata, T., Kawano, M., Igarashi, K., Yamato, I., and Kakinuma, Y. (2001) Catalytic properties of Na+-translocating V-ATPase in Enterococcus hirae, Biochim. Biophys. Acta, 1505, 75-81, doi: https://doi.org/10.1016/S0005-2728(00)00278-4.

    Article  CAS  PubMed  Google Scholar 

  19. Yokoyama, K., Nakano, M., Imamura, H., Yoshida, M., and Tamakoshi, M. (2003) Rotation of the proteolipid ring in the V-ATPase, J. Biol. Chem., 278, 24255-24258, doi: https://doi.org/10.1074/jbc.M303104200.

    Article  CAS  PubMed  Google Scholar 

  20. Corbalan-Garsia, S., Teruel, J. A., and Gomez-Fernandez, J. C. (1992) Characterization of Ruthenium Red-binding sites of the Ca2+-ATPase from sarcoplasmic reticulum and their interaction with Ca2+-binding sites, Biochemistry, 287, 767-774, doi: https://doi.org/10.1042/bj2870767.

    Article  Google Scholar 

  21. Wiangnon, K., Raksajit, W., and Incharoensakdi, A. (2007) Presence of a Na+-stimulated P-type ATPase in the plasma membrane of the alkaliphilic halotolerant cyanobacterium Aphanothece halophytica, FEMS Microbiol. Lett., 270, 139-145, doi: https://doi.org/10.1111/j.1574-6968.2007.00667.x.

    Article  CAS  PubMed  Google Scholar 

  22. Massyuk, N. P. (1973) Morphology, taxonomy, ecology and geographic distribution of the genus Dunaliella Teod. and prospects for its potential utilization, Naukova Dumka, Kiev, p. 244.

  23. Abdullaev, A. A., and Semenenko, V. E. (1974) Intensive cultivation and certain physiological characteristics of Dunaliella salina Teod., Soviet Plant Physiol., 21, 947-955.

    Google Scholar 

  24. Carter, S. G., and Karl, D. W. (1982) Inorganic phosphate assay with malachite green: an improvement and evaluation, J. Biochem. Biophys. Methods, 7, 7-13, doi: https://doi.org/10.1016/0165-022x(82)90031-8.

    Article  CAS  PubMed  Google Scholar 

  25. Simpson, I. A., and Sonne, O. (1982) A simple, rapid and sensitive method for measuring protein concentration in subcellular membrane fractions prepared by sucrose density ultracentrifugation, Anal. Biochem., 119, 424-427, doi: https://doi.org/10.1016/0003-2697(82)90608-x.

    Article  CAS  PubMed  Google Scholar 

  26. Krulwich, T. A. (1983) Na+/H+ antiporters, Biochim. Biophys. Acta, 726, 245-264, doi: https://doi.org/10.1016/0304-4173(83)90011-3.

    Article  CAS  PubMed  Google Scholar 

  27. Pagis, L. Ya., Popova, L. G., Andreev, I. M., and Balnokin, Yu. V. (2001) Ion specificity of Na+-transporting systems in the plasma membrane of the halotolerant alga Tetraselmis (Platymonas) viridis, Russ. J. Plant Physiol., 48, 281-286, doi: https://doi.org/10.1023/A:1016645829002.

    Article  CAS  Google Scholar 

  28. Kluge, C., and Dimroth, P. (1993) Specific protection by Na+ or Li+ of the F1Fo-ATPase of Propionigenium modestum from the reaction with dicyclohexylcarbodiimide, J. Biol. Chem., 268, 14557-14560.

    CAS  PubMed  Google Scholar 

  29. Popova, L. G., Shumkova, G. A., Andreev, I. M., and Balnokin, Yu. V. (2000) Na+-dependent electrogenic ATPase from the plasma membrane of the halotolerant microalga Dunaliella maritima, Doklady Biochemistry, 375, 235-238, doi: https://doi.org/10.1023/A:1026675923730.

    Article  CAS  PubMed  Google Scholar 

  30. Sussman, M. R., and Slayman, C. W. (1983) Modification of the Neurospora crassa plasma membrane H+-ATPase with N,N′-dicyclohexylcarbodiimide, J. Biol. Chem., 258, 1839-1843.

    CAS  PubMed  Google Scholar 

  31. Kinsella, J. L., Wehrle, J., Wilkins, N., and Sacktor, B. (1987) Inhibition of Na+-H+-exchange by N,N′-dicyclohexylcarbodiimide in isolated rat renal brush border membrane vesicles, J. Biol. Chem., 262, 7092-7097.

    CAS  PubMed  Google Scholar 

  32. Murakami, N., and Konishi, T. (1989) Mechanism of function of dicyclohexylcarbodiimide-sensitive Na+/H+-antiporter in Halobacterium halobium: pH effect, Arch. Biochem. Biophys., 271, 515-523, doi: https://doi.org/10.1016/0003-9861(89)90303-2.

    Article  CAS  PubMed  Google Scholar 

Download references

Funding

This work was financially supported by the Russian Foundation for Basic Research (project No. 20-04-00903).

Author information

Authors and Affiliations

  1. Timiryazev Institute of Plant Physiology, Russian Academy of Sciences, 127276, Moscow, Russia

    L. G. Popova, D. A. Matalin & Yu. V. Balnokin

Authors
  1. L. G. Popova
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. D. A. Matalin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Yu. V. Balnokin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to L. G. Popova.

Ethics declarations

The authors declare no conflict of interests in financial or any other sphere. This article does not contain any studies with human participants or animals performed by any of the authors.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Popova, L.G., Matalin, D.A. & Balnokin, Y.V. Electrogenesis in Plasma Membrane Fraction of Halotolerant Microalga Dunaliella maritima and Effects of N,N′-Dicyclohexylcarbodiimide. Biochemistry Moscow 85, 930–937 (2020). https://doi.org/10.1134/S0006297920080088

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1134/S0006297920080088

Keywords

Search

Navigation