Photosynthesis Research

, Volume 63, Issue 3, pp 225–236 | Cite as

Targeted inactivation of the psaK1, psaK2 and psaM genes encoding subunits of Photosystem I in the cyanobacterium Synechocystis sp. PCC 6803

  • Sushma Naithani
  • Jian-Min Hou
  • Parag R. Chitnis

Abstract

PsaK and PsaM are small, integral membrane proteins, which are associated with the Photosystem I complexes of cyanobacteria. The complete genome sequence of Synechocystis sp. PCC 6803 has revealed the presence of two unlinked psaK genes: psaK1 (ssr0390) and psaK2(sll0629). To investigate structural and functional roles of the PsaK1, PsaK2 and PsaM polypeptides in Synechocystis sp. PCC 6803, we generated targeted mutants that lack the functional psaK1, psaK2 or psaM genes. Inactivation of psaK1, psaK2 or psaM did not affect photoautotrophic growth, photosynthetic activity and accumulation of other subunits of the Photosystem I complex. The psaK1, psaK2 and psaK1psaK2 mutants showed normal levels of Photosystem I trimers, whereas the lack of PsaM resulted in a 75% reduction in the recovery of trimers compared to the wild type. A 6.2 kDa polypeptide was observed in the Photosystem I preparations from the wild type, but not from the psaK2 strain, suggesting the presence of PsaK2 in the Photosystem I complexes. Using reverse-transcription and polymerase chain reaction, we confirmed the expression of the psaK2 gene in Synechocystis sp. PCC 6803. To conclude, both psaK1 and psaK2 are expressed in Synechocystis sp. PCC 6803 and the absence of both proteins results in only a small reduction in Photosystem I electron transport. The PsaM subunit is required for the formation of stable Photosystem I trimers.

cyanobacteria Photosystem I PsaK PsaM targeted-mutagenesis 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Arnon D. 1949. Copper enzyme in isolated chloroplasts. Polyphenol oxidase in Beta vulgaris. Plant Physiol. 24: 1–14.PubMedGoogle Scholar
  2. Brettel K. 1997. Electron transfer and arrangement of the redox cofactors in Photosystem. I. Biochim. Biophys. Acta 1318: 322–373.CrossRefGoogle Scholar
  3. Chitnis P.R. 1996. Photosystem I. Plant Physiol. 111: 661–669.PubMedCrossRefGoogle Scholar
  4. Chitnis V.P. and Chitnis P.R. 1993. PsaL subunit is required for the formation of Photosystem I trimers in the cyanobacterium Synechocystis sp. PCC 6803. FEBS Lett. 336: 330–4.PubMedCrossRefGoogle Scholar
  5. Haehnel W., Jansen T., Gause K., Klosgen R.B., Stahl B., Michl D. et al. 1994. Electron transfer from plastocyanin to Photosystem I. EMBO J. 13: 1028–38.PubMedGoogle Scholar
  6. Golbeck J.H. 1994. Photosystem I in Cyanobacteria:. In: Bryant D.A. (ed.), The Molecular Biology of Cyanobacteria. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp. 179–220.Google Scholar
  7. Golden S.S., Brusslan J. and Haselkorn R. 1987. Genetic engineering of the cyanobacterial chromosome. Meth. Enzymol. 153: 215–231.PubMedGoogle Scholar
  8. Jansson S., Anderson B. and Scheller H.V. 1996. Nearest neighbour analysis of higher plant Photosystem I holocomplex. Plant Physiol. 112: 409–420.PubMedCrossRefGoogle Scholar
  9. Kaneko T., Sato S., Kotani H., Tanaka A., Asamizu E., Nakamura Y. et al. 1996. Sequence analysis of the genome of the unicellular Cyanobacterium Synechocystis sp. strain PCC 6803. II. Sequence determination of the entire genome and assignment of potential protein-coding regions. DNA Res. 3: 109–136.PubMedCrossRefGoogle Scholar
  10. Kjærulff S., Andersen B., Nielsen V.S., Moller B.L. and Okkels J.S. 1993. The PS I-K Subunit of Photosystem I from Barley (Hordeum vulgare L.): Evidence for a gene duplication of an ancestral PS I-G/K gene. J. Biol. Chem. 268: 18912–18916.PubMedGoogle Scholar
  11. Krauß N., Schubert W.-D., Klukas O., Fromme P., Witt H.T. and Saenger W. 1996. Photosystem I at 4 Å resolution represents the first structural model of a joint photosynthetic reaction centre and core antenna system. Nature Struct. Biol. 3: 965–973.PubMedCrossRefGoogle Scholar
  12. Manna P. and Chitnis P.R. 1999. Function and molecular genetics of Photosystem I. In: Singhal G.S., Renger G. and Sapory S.K. (eds), Concepts in Photobiology: Photosynthesis and Photomorphogenesis., pp. 212–251.Google Scholar
  13. Muhlenhoff U. and Chauvat F. 1996. Gene transfer and manipulation in the thermophillic cyanobacterium Synechococcus elongatus. Mol. Gen. Genet. 252: 93–100.PubMedCrossRefGoogle Scholar
  14. Nakamoto H. and Hasegawa M. 1999. Targeted inactivation of the gene psaK encoding a subunit of Photosystem I from the cyanobacterium Synechocystis sp. PCC 6803. Plant Cell. Physiol. 40: 9–16.PubMedGoogle Scholar
  15. Oh-oka H., Takahashi Y., Kuriyama K., Saeki K. and Matsubara H. 1988. The protein responsible for center A/B in spinach Photosystem I: isolation with iron sulfur cluster(s) and complete sequence analysis. J. Biochem. 103: 962–968.PubMedGoogle Scholar
  16. Okkels J.S., Nielsen S., Scheller H.V. and Møller B.L. 1992. A cDNA clone from barley encoding the precursor from the Photosystem I polypeptide PS I-G: Sequence similarity to PS I-K. Plant Mol. Biol. 18: 989–994.PubMedCrossRefGoogle Scholar
  17. Rippka R., Deruelles J., Waterbury J.B., Herdman M. and Stanier R.Y. 1979. Generic assignments, strain histories and properties of pure cultures of cyanobacteria. J. Gen. Microbiol. 111: 1–61.Google Scholar
  18. Rousseau F., Setif P. and Lagoutte B. 1993. Evidence for the involvement of PS I-E subunit in the reduction of ferredoxin by Photosystem I. EMBO J. 12: 1755–1765.PubMedGoogle Scholar
  19. Sambrook J., Fritsch E.F. and Maniatis T. 1989. Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York.Google Scholar
  20. Soukoulis V., Savikhin, Xu W., Chitnis P.R. and Struve W.S. 1999. Electronic Spectra of PS I mutants: the peripheral subunits do not bind red chlorophyll in Synechocystis sp. PCC 6803. Biophys J. 76: 2711–2715.PubMedCrossRefGoogle Scholar
  21. Sun J., Ke A., Jin P., Chitnis V.P. and Chitnis P.R. 1998. Isolation and functional study of Photosystem I subunits in the cyanobacterium Synechocystis sp. PCC 6803. Meth. Enzymol. 297: 124–139.PubMedGoogle Scholar
  22. Williams J.G.K. 1988. Construction of specific mutations in Photosystem II photosynthetic reaction center by genetic engineering methods in Synechocystis 6803. Meth. Enzymol. 167: 766–778.CrossRefGoogle Scholar
  23. Wynn R.M. and Malkin R. 1988a. Characterization of an isolated chloroplast membrane Fe-S protein and its identification as the Photosystem I Fe-SA/Fe-SB binding protein. FEBS Lett. 229: 293–297.CrossRefGoogle Scholar
  24. Wynn R.M. and Malkin R. 1988b. Interaction of plastocyanin with Photosystem I: a chemical crosslinking study of the polypeptide that binds plastocyanin. Biochemistry 27: 5863–9.PubMedCrossRefGoogle Scholar
  25. Xu Q., Armbrust T.S., Guikema J.A. and Chitnis P.R. 1994a. Organization of Photosystem I polypeptides: a structural interactions between PsaD and PsaL subunits. Plant Physiol. 106: 1057–1063.PubMedCrossRefGoogle Scholar
  26. Xu Q., Jung Y.S., Chitnis V.P., Guikema J.A., Golbeck J.H. and Chitnis P.R. 1994b. Mutational analysis of Photosystem I poly-peptides in Synechocystis sp. PCC 6803. Subunit requirements for reduction of NADP+ mediated by ferredoxin and flavodoxin. J. Biol. Chem. 2693: 21512–21518.Google Scholar
  27. Xu Q., Odom W.R., Guikema J.A., Chitnis V.P. and Chitnis P.R. 1994c. Targeted deletion of pasJ from the cyanobacterium Synechocystis sp. PCC 6803 indicates structural interactions between the PsaJ and PsaF subunits of Photosystem I. Plant Mol. Biol. 2624: 291–302.CrossRefGoogle Scholar
  28. Xu Q., Yu L., Chitnis V.P. and Chitnis P.R. 1994d. Function and organization of Photosystem I in a cyanobacterial mutant strain that lacks PsaF and PsaJ subunits. J. Biol. Chem. 269: 3205–11.PubMedGoogle Scholar
  29. Xu Q., Hoppe D., Chitnis V.P., Odom W.R., Guikema J.A. and Chitnis P.R. 1995. Mutational analysis of Photosystem I polypeptides in the cyanobacterium Synechocystis sp. PCC 6803. Targeted inactivation of psaI reveals the function of PsaI in the structural organization of PsaL. J. Biol. Chem. 270: 16243–16250.PubMedCrossRefGoogle Scholar
  30. Zilber A. and Malkin R. 1988. Ferredoxin cross-links to a 22 kD subunit of Photosystem I. Plant Physiol. 88: 810–814.PubMedGoogle Scholar

Copyright information

© Kluwer Academic Publishers 2000

Authors and Affiliations

  • Sushma Naithani
    • 1
  • Jian-Min Hou
    • 2
  • Parag R. Chitnis
    • 3
  1. 1.Department of Biochemistry, Biophysics and Molecular BiologyIowa State UniversityAmesUSA
  2. 2.Laboratory ofPhotosynthesis Basic Research, Institute of BotanyChinese Academy of SciencesBeijingPeoples' Republic of China
  3. 3.Department of Biochemistry, Biophysics and Molecular BiologyIowa State UniversityAmesUSA

Personalised recommendations