Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1988 Oct;85(19):7226–7230. doi: 10.1073/pnas.85.19.7226

Directed mutations affecting spectroscopic and electron transfer properties of the primary donor in the photosynthetic reaction center

Edward J Bylina 1, Douglas C Youvan 1,*
PMCID: PMC282157  PMID: 16578836

Abstract

Oligonucleotide-mediated mutagenesis has been used to change the histidine residues that act as axial ligands to the central Mg2+ ions of the “special pair” bacteriochlorophylls in the reaction center of Rhodobacter capsulatus. Histidine-173 of the L subunit has been replaced with glutamine, while histidine-200 of the M subunit has been replaced with glutamine, leucine, or phenylalanine. When leucine or phenylalanine is introduced at M200, one of the special pair bacteriochlorophylls is converted to bacteriopheophytin, which generates a heterodimer at the special pair binding site. The pigment composition of the reaction center is unaltered when either histidine is replaced with glutamine. All of these mutant reaction centers are photochemically active, although the electron transfer properties of heterodimer-containing reaction centers are altered. These mutations begin to define the structural parameters that determine whether bacteriochlorophyll or bacteriopheophytin will be incorporated into the tetrapyrrole binding sites of the photosynthetic reaction center. Our results demonstrate that the properties of the photosynthetic reaction center can be changed by directed mutagenesis, which makes this complex an excellent model for testing theories of electron transfer in biological systems.

Keywords: protein engineering, pigment composition, heterodimer, Rhodobacter capsulatus

Full text

PDF
7226

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Allen J. P., Feher G., Yeates T. O., Rees D. C., Deisenhofer J., Michel H., Huber R. Structural homology of reaction centers from Rhodopseudomonas sphaeroides and Rhodopseudomonas viridis as determined by x-ray diffraction. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8589–8593. doi: 10.1073/pnas.83.22.8589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blankenship R. E., Feick R., Bruce B. D., Kirmaier C., Holten D., Fuller R. C. Primary photochemistry in the facultative green photosynthetic bacterium Chloroflexus aurantiacus. J Cell Biochem. 1983;22(4):251–261. doi: 10.1002/jcb.240220407. [DOI] [PubMed] [Google Scholar]
  3. Bylina E. J., Ismail S., Youvan D. C. Plasmid pU29, a vehicle for mutagenesis of the photosynthetic puf operon in Rhodopseudomonas capsulata. Plasmid. 1986 Nov;16(3):175–181. doi: 10.1016/0147-619x(86)90055-7. [DOI] [PubMed] [Google Scholar]
  4. Chang C. H., Tiede D., Tang J., Smith U., Norris J., Schiffer M. Structure of Rhodopseudomonas sphaeroides R-26 reaction center. FEBS Lett. 1986 Sep 1;205(1):82–86. doi: 10.1016/0014-5793(86)80870-5. [DOI] [PubMed] [Google Scholar]
  5. Deisenhofer J., Epp O., Miki K., Huber R., Michel H. X-ray structure analysis of a membrane protein complex. Electron density map at 3 A resolution and a model of the chromophores of the photosynthetic reaction center from Rhodopseudomonas viridis. J Mol Biol. 1984 Dec 5;180(2):385–398. doi: 10.1016/s0022-2836(84)80011-x. [DOI] [PubMed] [Google Scholar]
  6. Ditta G., Stanfield S., Corbin D., Helinski D. R. Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank of Rhizobium meliloti. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7347–7351. doi: 10.1073/pnas.77.12.7347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Evans T. A., Katz J. J. Evidence for 5- and 6-coordinated magnesium in bacterio-chlorophyll a from visible absorption spectroscopy. Biochim Biophys Acta. 1975 Sep 8;396(3):414–426. doi: 10.1016/0005-2728(75)90147-4. [DOI] [PubMed] [Google Scholar]
  8. Fajer J., Brune D. C., Davis M. S., Forman A., Spaulding L. D. Primary charge separation in bacterial photosynthesis: oxidized chlorophylls and reduced pheophytin. Proc Natl Acad Sci U S A. 1975 Dec;72(12):4956–4960. doi: 10.1073/pnas.72.12.4956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Michel H., Epp O., Deisenhofer J. Pigment-protein interactions in the photosynthetic reaction centre from Rhodopseudomonas viridis. EMBO J. 1986 Oct;5(10):2445–2451. doi: 10.1002/j.1460-2075.1986.tb04520.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Nanba O., Satoh K. Isolation of a photosystem II reaction center consisting of D-1 and D-2 polypeptides and cytochrome b-559. Proc Natl Acad Sci U S A. 1987 Jan;84(1):109–112. doi: 10.1073/pnas.84.1.109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Norris J. R., Scheer H., Druyan M. E., Katz J. J. An electron-nuclear double resonance (ENDOR) study of the special pair model for photo-reactive chlorophyll in photosynthesis. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4897–4900. doi: 10.1073/pnas.71.12.4897. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Norris J. R., Uphaus R. A., Crespi H. L., Katz J. J. Electron spin resonance of chlorophyll and the origin of signal I in photosynthesis. Proc Natl Acad Sci U S A. 1971 Mar;68(3):625–628. doi: 10.1073/pnas.68.3.625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Scolnik P. A., Marrs B. L. Genetic research with photosynthetic bacteria. Annu Rev Microbiol. 1987;41:703–726. doi: 10.1146/annurev.mi.41.100187.003415. [DOI] [PubMed] [Google Scholar]
  14. Straley S. C., Parson W. W., Mauzerall D. C., Clayton R. K. Pigment content and molar extinction coefficients of photochemical reaction centers from Rhodopseudomonas spheroides. Biochim Biophys Acta. 1973 Jun 28;305(3):597–609. doi: 10.1016/0005-2728(73)90079-0. [DOI] [PubMed] [Google Scholar]
  15. Van der Rest M., Gingras G. The pigment complement of the photosynthetic reaction center isolated from Rhodospirillum rubrum. J Biol Chem. 1974 Oct 25;249(20):6446–6453. [PubMed] [Google Scholar]
  16. Williams J. C., Steiner L. A., Feher G. Primary structure of the reaction center from Rhodopseudomonas sphaeroides. Proteins. 1986 Dec;1(4):312–325. doi: 10.1002/prot.340010405. [DOI] [PubMed] [Google Scholar]
  17. Yen H. C., Marrs B. Growth of Rhodopseudomonas capsulata under anaerobic dark conditions with dimethyl sulfoxide. Arch Biochem Biophys. 1977 Jun;181(2):411–418. doi: 10.1016/0003-9861(77)90246-6. [DOI] [PubMed] [Google Scholar]
  18. Youvan D. C., Ismail S., Bylina E. J. Chromosomal deletion and plasmid complementation of the photosynthetic reaction center and light-harvesting genes from Rhodopseudomonas capsulata. Gene. 1985;38(1-3):19–30. doi: 10.1016/0378-1119(85)90199-4. [DOI] [PubMed] [Google Scholar]
  19. Youvan D. C., Marrs B. L. Molecular genetics and the light reactions of photosynthesis. Cell. 1984 Nov;39(1):1–3. doi: 10.1016/0092-8674(84)90185-5. [DOI] [PubMed] [Google Scholar]
  20. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis: a simple method using two oligonucleotide primers and a single-stranded DNA template. DNA. 1984 Dec;3(6):479–488. doi: 10.1089/dna.1.1984.3.479. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES