Advertisement

Photosynthesis Research

, Volume 37, Issue 2, pp 139–146 | Cite as

Cloning and sequencing of mutantpsbB genes of the cyanobacteriumSynechocystis PCC 6803

  • Swetlana Yu. Ermakova
  • Irina V. Elanskaya
  • Kai-Uwe Kallies
  • Andreas Weihe
  • Thomas Börner
  • Sergey V. Shestakov
Regular Paper

Abstract

Ten strains from a collection of mutants ofSynechocystis 6803 defective in Photosystem II (PS II) function were transformed with chromosomal DNA of wild-type and mutant cells. Cross hybridization data allowed to identify four groups of PS II-mutants. Highly efficient transformation was observed between different mutant groups, but not within the groups. Restoration of photosynthetic activity of the mutant cells was also achieved by transformation with different parts of a 5.6 kbBam HI fragment of wild typeSynechocystis DNA containing thepsbB gene. Each group of mutants was transformed to photoautotrophic growth by specific subfragments of thepsbB gene. DNA fragments of four selected mutant strains hybridizing with thepsbB gene were isolated and sequenced. The mutations were identified as a single nucleotide insertion or substitution leading to stop codon formation in two of the mutants, as a deletion of 12 nucleotides, or as a nucleotide substitution resulting in an amino acid substitution in the other two mutants. Deletion of 12 nucleotides in mutant strain PMB1 and stop codon formation in strain NF16 affect membrane-spanning regions of the gene product, the CP 47 protein.

Key words

chlorophyll-binding protein CP47 DNA sequence gene analysis mutagenesis Photosystem II 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Barsky EL, Gubanova ON, Elanskaya IV, Samuilov VD, Stanbekova GE and Shestakov SV (1989) Mutants of cyanobacteriumSynechocystis 6803 with defects in electron transport on the donor side of Photosystem II. Biologicheskye Nauki 7: 29–33 (in Russian)Google Scholar
  2. Bicker TM (1990) The structure and function of CPa-1 and CPa-2 in Photosystem II. Photosynth Res 24: 1–13Google Scholar
  3. Bukharov AA, Kolosov VL and Zolotarev AS (1988) Nucleotide sequence of rye chloroplast DNA fragment encodingpsbB andpsbH genes. Nucleic Acids Res 16: 8737PubMedGoogle Scholar
  4. Carpenter SD and Vermaas WFJ (1989) Directed mutagenesis to probe the structure and function of Photosystem II. Physiol Plant 77: 436–443Google Scholar
  5. Eatom-Rye JJ and Vermaas WFJ (1991) Oligonucleotide-directed mutagenesis ofpsbB, the gene encoding CP 47, employing a deletion mutant strain of the cyanobacteriumSynechocystis sp. PCC 6803. Plant Mol Biol 17: 1165–1177PubMedGoogle Scholar
  6. Grigorieva G and Shestakov S (1982) Transformation in the cyanobacteriumSynechocystis 6803. FEMS Microbiol Lett 13: 367–370Google Scholar
  7. Gulyaev BA and Tetenin VL (1983) The native criteria of pigment-protein complexes and the features of their organization in vivo. Proc Acad Sci USSR Biol Series 4: 536–552 (in Russian)Google Scholar
  8. Ikeuchi M and Inoue Y (1988) A new Photosystem II reaction center component (4.8 kDa protein) encoded by chloroplast genome. FEBS Lett 241: 99–104PubMedGoogle Scholar
  9. Keller M, Weil JH and Nair CKK (1989) Nucleotide sequence of thepsbB gene ofEuglena gracilis. Plant Mol Biol 13: 723–725PubMedGoogle Scholar
  10. Lang JD and Haselkorn R (1989) Isolation, sequence and transcription of the gene encoding the Photosystem II chlorophyll-binding protein, CP-47, in the cyanobacteriumAnabaena 7120. Plant Mol Biol 13: 441–456PubMedGoogle Scholar
  11. Lehmbek J, Stummann BM and Henningsen KW (1989) Nucleotide sequence of the 5.6 kbppsbB operon of pea chloroplast DNA. Physiol Plant 76: 57–64Google Scholar
  12. Lyadsky VV, Gorbunov MYu and Venediktov PS (1987) Impulse fluorometer for study of the primary photosynthetic reactions in green plants. Biologicheskye Nauki 12: 96–100 (in Russian)Google Scholar
  13. Maniatis T, Fritsch EF and Sambrook J (1982) Molecular Cloning: A Laboratory Manual. Cold Spring Harbor: Cold Spring Harbor PressGoogle Scholar
  14. Morris J and Herrmann RG (1984) Nucleotide sequence of the gene for the chlorophylla apoprotein of the Photosystem II reaction center from spinach. Nucleic Acids Res 12: 2837–2850PubMedGoogle Scholar
  15. Pakrasi HB, Williams JGK and Arntzen CJ (1987) Genetically engineered cytochromeb559 mutants of the cyano-bacteriumSynechocystis 6803. In: Biggins J (ed) Progress in Photosynthesis Research, Vol 4, pp 813–816. Kluwer Academic Publishers, Dordrecht.Google Scholar
  16. Rippka R, Deruelles J, Waterbury JB, Herdmann M and Stanier RT (1979) Generic assignments, strain histories and properties of pure cultures of cyanobacteria. J Gen Microbiol 111: 1–61Google Scholar
  17. Rock CD, Barkan A and Taylor WC (1987) The maize plastidpsbB-psbF-petB-petD gene cluster: Spliced and unsplicedpetB andpetD RNAs encode alternative products. Curr Genet 12: 69–77PubMedGoogle Scholar
  18. Sanger F, Nicklen S and Coulsen AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467PubMedGoogle Scholar
  19. Shestakov SV, Koksharova OA, Groshev VV and Elanskaya IV (1988) Photosynthesis-defective mutants of the cyano-bacteriumSynechocystis 6803. Biologicheskye nauki 1: 75–80 (in Russian)Google Scholar
  20. Shestakov SV, Elanskaya IV, Ermakova SY, Andrianov VM and Ulmasov TN (1989) Cloning of the DNA fragments transforming to photoautotrophic growth photosynthesis-defective mutants of the cyanobacteriumSynechocystis 6803. Dokl Akad nauk SSSR 308: 211–214 (in Russian)Google Scholar
  21. Vermaas WFJ, Ikeuchi M and Inoue Y (1988) Protein composition of the Photosystem II core complex in genetically engineered mutants of the cyanobacteriumSynechocystis sp. PCC 6803. Photosynth Res 17: 97–113Google Scholar
  22. Vermaas WFJ, Williams JGK and Arntzen CJ (1987) Sequencing and modification ofpsbB, the gene encoding the CP-47 protein of Photosystem II, in the cyanobacteriumSynechocystis 6803. Plant Mol Biol 8: 317–326Google Scholar
  23. Vermaas WFJ, Williams JGK, Rutherford AW, Mathis P and Arntzen CJ (1986) Genetically engineered mutant of the cyanobacteriumSynechocystis 6803 lacks the Photosystem II chlorophyll binding protein CP 47. Proc Natl Acad Sci USA 83: 9474–9477Google Scholar
  24. Williams JGK (1988) Construction of specific mutations in the Photosystem II photosynthetic reaction center by genetic engineering methods in the cyanobacteriumSynechocystis 6803. Methods Enzymol 167: 766–778Google Scholar
  25. Yanisch-Perron C, Vieira J and Messing J (1985) Improved M13 phage cloning vectors and host strains: Nucleotide sequence of the M13mp18 and pUC19 vectors. Gene 33: 103–119PubMedGoogle Scholar

Copyright information

© Kluwer Academic Publishers 1993

Authors and Affiliations

  • Swetlana Yu. Ermakova
    • 1
  • Irina V. Elanskaya
    • 1
  • Kai-Uwe Kallies
    • 2
  • Andreas Weihe
    • 2
  • Thomas Börner
    • 2
  • Sergey V. Shestakov
    • 1
  1. 1.Department of GeneticsMoscow State UniversityMoscowRussian Federation
  2. 2.Department of Biology, Institute of GeneticsHumboldt UniversityBerlinGermany

Personalised recommendations