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Plant Biol (Stuttg) 2004; 6(3): 358-368
DOI: 10.1055/s-2004-817893
Original Paper

Georg Thieme Verlag Stuttgart KG · New York

Hg2+ Reacts with Different Components of the NADPH : Protochlorophyllide Oxidoreductase Macrodomains

K. Solymosi1 , K. Lenti1 , B. Myśliwa-Kurdziel2 , J. Fidy3 , 4 , K. Strzałka2 , B. Böddi1
  • 1Department of Plant Anatomy, Eötvös University, Budapest, Hungary
  • 2Department of Plant Physiology and Biochemistry, Jagiellonian University, Krakow, Poland
  • 3Department of Biophysics and Radiation Biology, Semmelweis University, Budapest, Hungary
  • 4MTA-SE Research Group for Biophysics, Hungarian Academy of Science, Budapest, Hungary
Further Information

Publication History

Publication Date:
14 May 2004 (online)

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Abstract

The molecular background of Hg2+-induced inhibition of protochlorophyllide (Pchlide) photoreduction was investigated in homogenates of dark-grown wheat leaves. Our earlier work showed that 15 min incubation with 10-2 M Hg2+ completely inhibits the activity of NADPH : Pchlide oxidoreductase ([Lenti et al., 2002]). Detailed analysis of spectra recorded at 10 K indicated the appearance of emission bands at 638 and 650 nm, which are characteristic for NADP+-Pchlide complexes. Fluorescence emission spectra recorded with different excitation wavelengths, fluorescence lifetime measurements and the analysis of acetone extractions revealed that Hg2+ can also react directly with Pchlide, resulting in protopheophorbide formation. At 10-3 M Hg2+, the phototransformation was complete but the blue shift of the chlorophyllide emission band speeded up remarkably. This indicates oxidation of the NADPH molecules that have a structural role in keeping together the etioplast inner membrane components. We suggest a complex model for the Hg2+ effect: depending on concentration it can react with any components of the NADPH : Pchlide oxidoreductase macrodomains.

Key words

Mercury - Hg2+ - protochlorophyllide - NADPH - protopheophorbide - Shibata shift - activity loss

References

  • 1 Aronsson H., Sundqvist C., Timko M. P., Dahlin C.. The importance of the C-terminal region and Cys residues for the membrane association of the NADPH:protochlorophyllide oxidoreductase in pea.  FEBS Lett.. (2001);  502 11-15
    CrossrefPubMedGoogle Scholar
  • 2 Aylward G. H., Findlay T. J. V.. S. I. Chemical Data. Sydney; John Wiley and Sons (1974)
    Google Scholar
  • 3 Barlton D., Smith A. M.. Mercury binding to human haemoglobin.  Experientia. (1973);  29 1178-1179
    PubMedGoogle Scholar
  • 4 Böddi B.. Spectral, biochemical and structural changes connected to protochlorophyllide photoreduction in chlorophyll biosynthesis.  Human Envir. Sci.. (1994);  3 39-55
    PubMedGoogle Scholar
  • 5 Böddi B., Franck F.. Room temperature fluorescence spectra of protochlorophyllide and chlorophyllide forms in etiolated bean leaves.  J. Photochem. Photobiol.. (1997);  41 73-82
    PubMedGoogle Scholar
  • 6 Böddi B., Lindsten A., Ryberg M., Sundqvist C.. On the aggregational states of protochlorophyllide and its protein complexes in wheat etioplasts.  Physiol. Plant.. (1989);  76 135-143
    PubMedGoogle Scholar
  • 7 Böddi B., Oravecz A. R., Lehoczki E.. Effect of cadmium on organization and photoreduction of protochlorophyllide in dark-grown leaves and etioplast inner membrane preparations of wheat.  Photosynthetica. (1995);  31 411-420
    PubMedGoogle Scholar
  • 8 Böddi B., Popovic R., Franck F.. Early reactions of light-induced protochlorophyllide and chlorophyllide transformations analyzed in vivo at room temperature with a diode array spectrofluorometer.  J. Photochem. Photobiol. B: Biol.. (2003);  69 31-39
    PubMedGoogle Scholar
  • 9 Böddi B., Ryberg M., Sundqvist C.. The formation of a short-wavelength chlorophyllide form at partial phototransformation of protochlorophyllide in etioplast inner membranes.  Photochem. Photobiol.. (1991);  53 667-673
    PubMedGoogle Scholar
  • 10 Böddi B., Ryberg M., Sundqvist C.. Identification of four universal protochlorophyllide forms in dark-grown leaves by analyses of the 77 K fluorescence emission spectra.  J. Photochem. Photobiol. B: Biol.. (1992);  12 389-401
    PubMedGoogle Scholar
  • 11 Böddi B., Ryberg M., Sundqvist C.. Analysis of the 77 K fluorescence excitation spectra of isolated etioplast inner membranes.  J. Photochem. Photobiol. B: Biol.. (1993);  21 125-133
    PubMedGoogle Scholar
  • 12 Boening D. W.. Ecological effects, transport, and fate of mercury: a general review.  Chemosphere. (2000);  40 1335-1351
    CrossrefPubMedGoogle Scholar
  • 13 Cho U. H., Park J. O.. Mercury-induced oxidative stress in tomato seedlings.  Plant Sci.. (2000);  156 1-9
    CrossrefPubMedGoogle Scholar
  • 14 Clijsters H., van Asche F.. Inhibition of photosynthesis by heavy metals.  Photosynth. Res.. (1985);  7 31-40
    CrossrefPubMedGoogle Scholar
  • 15 Dahlin C., Aronsson H., Wilks H. M., Lebedev N., Sundqvist C., Timko M. P.. The role of protein surface charge in catalytic activity and chloroplast membrane association of the pea NADPH-protochlorophyllide oxidoreductase as revealed by alanine scanning mutagenesis.  Plant. Mol. Biol.. (1999);  39 309-323
    CrossrefPubMedGoogle Scholar
  • 16 De Filippis L. F.. The effect of heavy metals on the absorption spectra of Chlorella cells and chlorophyll solutions.  Z. Pflanzenphysiol.. (1979);  93 129-137
    PubMedGoogle Scholar
  • 17 De Filippis L. F., Pallaghy C. K.. The effect of sub-lethal concentrations of mercury and zinc on Chlorella. II. Photosynthesis and pigment composition.  Z. Pflanzenphysiol.. (1976);  78 314-322
    PubMedGoogle Scholar
  • 18 De Filippis L. F., Hampp R., Ziegler H.. The effects of sublethal concentrations of zinc, cadmium and mercury on Euglena. Growth and pigments.  Z. Pflanzenphysiol.. (1981 a);  101 37-47
    PubMedGoogle Scholar
  • 19 De Filippis L. F., Hampp R., Ziegler H.. The effects of sublethal concentrations of zinc, cadmium and mercury on Euglena. II. Respiration, photosynthesis and photochemical activities.  Arch. Microbiol.. (1981 b);  128 407-411
    CrossrefPubMedGoogle Scholar
  • 20 El Hamouri B., Sironval C.. NADP+/NADPH control of the protochlorophyllide-, chlorophyllide-proteins in cucumber etioplasts. Photochem.  Photobiophys.. (1980);  1 219-223
    PubMedGoogle Scholar
  • 21 Eullaffroy P., Salvetat R., Franck F., Popovic R.. Temperature dependence of chlorophyll(ide) spectral shifts and photoactive protochlorophyllide regeneration after flash in etiolated barley leaves.  Photochem. Photobiol.. (1995);  62 751-756
    PubMedGoogle Scholar
  • 22 Foster T. J.. Plasmid determined resistance to antimicrobial drugs and toxic metal ions in bacteria.  Microbiol. Rev.. (1983);  47 361-409
    PubMedGoogle Scholar
  • 23 Franck F., Bereza B., Böddi B.. Protochlorophyllide-NADP+ and protochlorophyllide-NADPH complexes and their regeneration after flash illumination in leaves and etioplast membranes of dark-grown wheat.  Photosynth. Res.. (1999);  59 53-61
    CrossrefPubMedGoogle Scholar
  • 24 Funahashi S., Inada Y., Inamo M.. Dynamic study of metal-ion incorporation into porphyrins based on the dynamic characterization of metal ions and sitting-atop complex formation.  Anal. Sci.. (2001);  17 917-927
    CrossrefPubMedGoogle Scholar
  • 25 Goedheer J. C.. Visible absorption and fluorescence of chlorophyll and its aggregates in solution. Vernon, L. P. and Seely, G. R., eds. The Chlorophylls. New York, Boston; Academic Press (1966): 147-184
    Google Scholar
  • 26 Gratton E., Limkeman M.. A continuously variable frequency cross-correlation phase fluorometer with picosecond resolution.  Biophys. J.. (1983);  44 315-324
    PubMedGoogle Scholar
  • 27 Griffiths W. T.. Reconstitution of chlorophyll formation by isolated etiplast membranes.  Biochem. J.. (1978);  174 681-692
    PubMedGoogle Scholar
  • 28 Griffiths W. T.. Protochlorophyllide photoreduction. Scheer H., ed. Chlorophylls. Boca Raton, Florida; CRC Press Inc. (1991): 433-450
    Google Scholar
  • 29 Heyes D. J., Martin G. E. M., Reid R. J., Hunter C. N., Wilks H. M.. NADPH: protochlorophyllide oxidoreductase from Synechocystis: overexpression, purification and preliminary characterisation.  FEBS Lett.. (2000);  483 47-51
    CrossrefPubMedGoogle Scholar
  • 30 Hu Q., Yang G., Yin J., Yao Y.. Determination of trace lead, cadmium and mercury by on-line column enrichment followed by RP-HPLC as metal-tetra-(4-bromophenyl)-porphyrin chelates.  Talanta. (2002);  57 751-756
    PubMedGoogle Scholar
  • 31 Kis-Petik K., Böddi B., Kaposi A. D., Fidy J.. Protochlorophyllide forms and energy transfer in dark-grown wheat leaves. Studies by conventional and laser excited fluorescence spectroscopy between 10 K-100 K.  Photosynth. Res.. (1999);  60 87-98
    CrossrefPubMedGoogle Scholar
  • 32 Klein S., Schiff J. A.. The correlated appearance of prolamellar bodies, protochlorophyll(ide) species, and the Shibata shift during development of bean etioplasts in the dark.  Plant. Physiol.. (1972);  49 619-626
    PubMedGoogle Scholar
  • 33 Küpper H., Küpper F., Spiller M.. Environmental relevance of heavy metal-substituted chlorophylls using the example of water plants.  J. Exp. Bot.. (1996);  47 259-266
    PubMedGoogle Scholar
  • 34 Küpper H., Küpper F., Spiller M.. In situ detection of heavy metal substituted chlorophylls in water plants.  Photosynth. Res.. (1998);  58 123-133
    CrossrefPubMedGoogle Scholar
  • 35 Küpper H., Šetlik I., Spiller M., Küpper F. C., Prášil O.. Heavy metal-induced inhibition of photosynthesis: targets of in vivo heavy metal chlorophyll formation.  J. Phycol.. (2002);  38 429-441
    CrossrefPubMedGoogle Scholar
  • 36 Lakowicz J. R., Lacko G., Cherek H., Gratton E., Limkeman M.. Analysis of fluorescence decay kinetics from variable-frequency phase shift and modulation data.  Biophys. J.. (1984);  46 463-477
    PubMedGoogle Scholar
  • 37 Lau S., Sarkar B.. Inorganic mercury(II)-binding component in normal human blood serum.  J. Toxicol. Environ. Health. (1979);  5 907-916
    PubMedGoogle Scholar
  • 38 Lebedev N., Timko M. P.. Protochlorophyllide photoreduction.  Photosynth. Res.. (1998);  58 5-23
    CrossrefPubMedGoogle Scholar
  • 39 Lenti K., Fodor F., Böddi B.. Mercury inhibits the activity of the NADPH:protochlorophyllide oxidoreductase (POR).  Photosynthetica. (2002);  40 145-151
    CrossrefPubMedGoogle Scholar
  • 40 Magnaval R., Batti R., Thiessard J.. Methylmercury effect on rat liver mitochondrial deshydrogenases.  Experimentia. (1975);  31 406-407
    PubMedGoogle Scholar
  • 41 Myśliwa-Kurdziel B., Strzałka K.. Influence of metals on biosynthesis of photosynthetic pigments. Prasad, M. N. V. and Strzałka, K., eds. Physiology and Biochemistry of Metal Toxicity and Tolerance in Plants. Dordrecht, Boston, London; Kluwer Academic Publishers (2002): 201-227
    Google Scholar
  • 42 Myśliwa-Kurdziel B., Franck F., Strzałka K.. Analysis of fluorescence lifetime of protochlorophyllide and chlorophyllide in isolated etioplast membranes measured from multifrequency cross-correlation phase fluorometry.  Photochem. Photobiol.. (1999);  70 616-623
    CrossrefPubMedGoogle Scholar
  • 43 Ogawa M., Tsutsui Y., Konishi M.. Effects of illumination on absorption peak shifts in spectra of intact etiolated cotyledons of Pharbitis nil. II. Effects of leaf age on protochlorophyllide regeneration and the Shibata shift.  Plant Cell Physiol.. (1978);  19 127-132
    PubMedGoogle Scholar
  • 44 Oosawa N., Masuda T., Awai K., Fusada N., Shimada H., Ohto H., Takamiya K.. Identification and light-induced expression of a novel gene of NADPH:protochlorophyllide oxidoreductase isoform in Arabidopsis thaliana. .  FEBS Lett.. (2000);  474 113-136
    PubMedGoogle Scholar
  • 45 Ouazzani-Chahdi M. A., Schoefs B., Franck F.. Isolation and characterisation of photoactive complexes of NADPH:protochlorophyllide oxidoreductase from wheat.  Planta. (1998);  206 673-680
    CrossrefPubMedGoogle Scholar
  • 46 Patra M., Sharma A.. Mercury toxicity in plants.  Bot. Rev.. (2000);  66 379-422
    PubMedGoogle Scholar
  • 47 Pearson R. G.. Hard and soft acids and bases.  J. Am. Chem. Soc.. (1963);  85 3533-3539
    CrossrefPubMedGoogle Scholar
  • 48 Romanowska E.. Gas exchange functions in metal stressed plants. Prasad, M. N. V. and Strzałka, K., eds. Physiology and Biochemistry of Metal Toxicity and Tolerance in Plants. Dordrecht, Boston, London; Kluwer Academic Publishers (2002): 257-285
    Google Scholar
  • 49 Ryberg M., Dehesh K.. Localization of NADPH:protochlorophyllide oxidoreductase in dark-grown wheat (Triticum aestivum) by immuno-electron microscopy before and after transformation of the prolamellar bodies.  Physiol. Plant.. (1986);  66 616-624
    PubMedGoogle Scholar
  • 50 Ryberg M., Sundqvist C.. Spectral forms of protochlorophyllide in prolamellar bodies and prothylakoids fractionated from wheat etioplasts.  Physiol. Plant.. (1982);  56 133-138
    PubMedGoogle Scholar
  • 51 Ryberg M., Sundqvist C.. The regular ultrastructure of isolated prolamellar bodies depends on the presence of membrane-bound NADPH-protochlorophyllide oxidoreductase.  Physiol. Plant.. (1988);  73 218-226
    PubMedGoogle Scholar
  • 52 Savitzky A., Golay M. J. E.. Smoothing and differentiation of data by simplified least square procedures.  Anal. Chem.. (1964);  36 1627
    CrossrefPubMedGoogle Scholar
  • 53 Schlegel H., Douglas L., Hüttermann A.. Whole plant aspects of heavy metal induced changes in CO2 uptake and water relations of spruce (Picea abies) seedlings.  Physiol. Plant.. (1987);  69 265-270
    PubMedGoogle Scholar
  • 54 Shibata K.. Spectroscopic studies on chlorophyll formation in intact leaves.  J. Biochem.. (1957);  44 147-173
    PubMedGoogle Scholar
  • 55 Smeller L.. How precise are the positions of computer-determined peaks?.  Appl. Spectrosc.. (1998);  52 1623-1626
    CrossrefPubMedGoogle Scholar
  • 56 Sperling U., Franck F., van Cleve B., Frick G., Apel K., Armstrong G. A.. Etioplast differentation in Arabidopsis: both PORA and PORB restore the prolamellar body and photoactive protochlorophyllide-P655 to the cop1 photomorphogenic mutant.  Plant Cell. (1998);  10 283-296
    CrossrefPubMedGoogle Scholar
  • 57 Sundberg J., Ersson B., Lönnerdal B., Oskarsson A.. Protein binding of mercury in milk and plasma from mice and man - a comparison between methylmercury and inorganic mercury.  Toxicology. (1999);  137 169-184
    CrossrefPubMedGoogle Scholar
  • 58 Suszcyńsky E. M., Shann J. R.. Phytotoxicity and accumulation of mercury in tobacco subjected to different exposure routes.  Environ. Toxicol. Chem.. (1995);  14 61-67
    PubMedGoogle Scholar
  • 59 Tabata M.. Kinetic evidence for short-lived intermediates in metalloporphyrin formation.  J. Mol. Liquids. (1995);  65 - 66 221-228
    CrossrefPubMedGoogle Scholar
  • 60 Teakle G. R., Griffiths W. T.. Cloning, characterization and import studies on protochlorophyllide reductase from wheat (Triticum aestivum). .  Biochem. J.. (1993);  296 225-230
    PubMedGoogle Scholar
  • 61 Vallee B. L., Ulmer D. D.. Biochemical effects of mercury, cadmium and lead.  Ann. Rev. Biochem.. (1972);  41 91-128
    PubMedGoogle Scholar
  • 62 Van Bochove A. C., Griffiths W. T., Van Grondelle R.. The primary reaction in the photoreduction of protochlorophyllide monitored by nanosecond fluorescence measurements.  Photochem. Photobiol.. (1984);  39 101-106
    PubMedGoogle Scholar
  • 63 Wiktorsson B., Engdahl S., Zhong L. B., Böddi B., Ryberg M., Sundqvist C.. The effect of cross-linking of the subunits of NADPH:protochlorophyllide oxidoreductase on the aggregational state of protochlorophyllide.  Photosynthetica. (1993);  29 205-218
    PubMedGoogle Scholar
  • 64 Wiktorsson B. M., Ryberg M., Gough S., Sundqvist C.. Isoelectric focusing of pigment-protein complexes solubilized from non-irradiated and irradiated prolamellar bodies.  Physiol. Plant.. (1992);  85 659-669
    CrossrefPubMedGoogle Scholar
  • 65 Zalups R. K., Barfuss D. W.. Nephrotoxicity of inorganic mercury co-administered with L-cysteine.  Toxicology. (1996);  109 15-29
    CrossrefPubMedGoogle Scholar

B. Böddi

Department of Plant Anatomy
Eötvös University

Pázmány P. sétány 1/C

Budapest H-1117

Hungary

Email: bbfotos@ludens.elte.hu

Guest Editor: F. Loreto

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