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. 1993 Aug;175(16):5066–5077. doi: 10.1128/jb.175.16.5066-5077.1993

Complementation analysis and regulation of CO2 fixation gene expression in a ribulose 1,5-bisphosphate carboxylase-oxygenase deletion strain of Rhodospirillum rubrum.

D L Falcone 1, F R Tabita 1
PMCID: PMC204973  PMID: 8349547

Abstract

A ribulose 1,5-bisphosphate carboxylase-oxygenase (RubisCO) deletion strain of Rhodospirillum rubrum that was incapable of photolithoautotrophic growth was constructed. Photoheterotrophic growth, however, was possible for the R. rubrum RubisCO deletion strain when oxidized carbon compounds such as malate were supplied. The R. rubrum RubisCO-deficient strain was not complemented to photolithoautotrophic growth by various R. rubrum DNA fragments that contain the gene encoding RubisCO, cbbM. When the R. rubrum cbbM deletion strain harbored plasmids containing R. rubrum DNA inserts with at least 2.0 kb preceding the translational start site of the cbbM gene, RubisCO activity and RubisCO antigen were detected. Lack of RubisCO expression was therefore not the cause for the failure to complement the cbbM mutant strain. Interestingly, DNA fragments encoding either of two complete Calvin-Benson-Bassham CO2- fixation (cbb) gene operons from Rhodobacter sphaeroides were able to complement the R. rubrum RubisCO deletion strain to photolithoautotrophic growth. The same R. rubrum DNA fragments that failed to complement the R. rubrum cbbM deletion strain successfully complemented the RubisCO deletion strain of R. sphaeroides, pointing to distinct differences in the regulation of metabolism and the genetics of photolithoautotrophic growth in these two organisms. A number of cbb genes were identified by nucleotide sequence analysis of the region upstream of cbbM. Included among these was an open reading frame encoding a cbbR gene showing a high degree of sequence similarity to known lysR-type CO2 fixation transcriptional activator genes. The placement and orientation of the cbbR transcriptional regulator gene in R. rubrum are unique.

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Selected References

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  1. Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
  2. Buchanan B. B., Evans M. C., Arnon D. I. Ferredoxin-dependent carbon assimilation in Rhodospirillum rubrum. Arch Mikrobiol. 1967;59(1):32–40. doi: 10.1007/BF00406314. [DOI] [PubMed] [Google Scholar]
  3. Chen J. H., Gibson J. L., McCue L. A., Tabita F. R. Identification, expression, and deduced primary structure of transketolase and other enzymes encoded within the form II CO2 fixation operon of Rhodobacter sphaeroides. J Biol Chem. 1991 Oct 25;266(30):20447–20452. [PubMed] [Google Scholar]
  4. Ditta G., Schmidhauser T., Yakobson E., Lu P., Liang X. W., Finlay D. R., Guiney D., Helinski D. R. Plasmids related to the broad host range vector, pRK290, useful for gene cloning and for monitoring gene expression. Plasmid. 1985 Mar;13(2):149–153. doi: 10.1016/0147-619x(85)90068-x. [DOI] [PubMed] [Google Scholar]
  5. Falcone D. L., Quivey R. G., Jr, Tabita F. R. Transposon mutagenesis and physiological analysis of strains containing inactivated form I and form II ribulose bisphosphate carboxylase/oxygenase genes in Rhodobacter sphaeroides. J Bacteriol. 1988 Jan;170(1):5–11. doi: 10.1128/jb.170.1.5-11.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Falcone D. L., Tabita F. R. Expression of endogenous and foreign ribulose 1,5-bisphosphate carboxylase-oxygenase (RubisCO) genes in a RubisCO deletion mutant of Rhodobacter sphaeroides. J Bacteriol. 1991 Mar;173(6):2099–2108. doi: 10.1128/jb.173.6.2099-2108.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Figurski D. H., Helinski D. R. Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1648–1652. doi: 10.1073/pnas.76.4.1648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Figurski D., Meyer R., Miller D. S., Helinski D. R. Generation in vitro of deletions in the broad host range plasmid RK2 using phage Mu insertions and a restriction endonuclease. Gene. 1976;1(1):107–119. doi: 10.1016/0378-1119(76)90010-x. [DOI] [PubMed] [Google Scholar]
  9. Fisher R. F., Egelhoff T. T., Mulligan J. T., Long S. R. Specific binding of proteins from Rhizobium meliloti cell-free extracts containing NodD to DNA sequences upstream of inducible nodulation genes. Genes Dev. 1988 Mar;2(3):282–293. doi: 10.1101/gad.2.3.282. [DOI] [PubMed] [Google Scholar]
  10. Fisher R. F., Long S. R. DNA footprint analysis of the transcriptional activator proteins NodD1 and NodD3 on inducible nod gene promoters. J Bacteriol. 1989 Oct;171(10):5492–5502. doi: 10.1128/jb.171.10.5492-5502.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gibson J. L., Chen J. H., Tower P. A., Tabita F. R. The form II fructose 1,6-bisphosphatase and phosphoribulokinase genes form part of a large operon in Rhodobacter sphaeroides: primary structure and insertional mutagenesis analysis. Biochemistry. 1990 Sep 4;29(35):8085–8093. doi: 10.1021/bi00487a014. [DOI] [PubMed] [Google Scholar]
  12. Gibson J. L., Falcone D. L., Tabita F. R. Nucleotide sequence, transcriptional analysis, and expression of genes encoded within the form I CO2 fixation operon of Rhodobacter sphaeroides. J Biol Chem. 1991 Aug 5;266(22):14646–14653. [PubMed] [Google Scholar]
  13. Gibson J. L., Tabita F. R. Isolation of the Rhodopseudomonas sphaeroides form I ribulose 1,5-bisphosphate carboxylase/oxygenase large and small subunit genes and expression of the active hexadecameric enzyme in Escherichia coli. Gene. 1986;44(2-3):271–278. doi: 10.1016/0378-1119(86)90191-5. [DOI] [PubMed] [Google Scholar]
  14. Gibson J. L., Tabita F. R. Localization and mapping of CO2 fixation genes within two gene clusters in Rhodobacter sphaeroides. J Bacteriol. 1988 May;170(5):2153–2158. doi: 10.1128/jb.170.5.2153-2158.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gibson J. L., Tabita F. R. Organization of phosphoribulokinase and ribulose bisphosphate carboxylase/oxygenase genes in Rhodopseudomonas (Rhodobacter) sphaeroides. J Bacteriol. 1987 Aug;169(8):3685–3690. doi: 10.1128/jb.169.8.3685-3690.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Goethals K., Van Montagu M., Holsters M. Conserved motifs in a divergent nod box of Azorhizobium caulinodans ORS571 reveal a common structure in promoters regulated by LysR-type proteins. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1646–1650. doi: 10.1073/pnas.89.5.1646. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hallenbeck P. L., Kaplan S. Cloning of the gene for phosphoribulokinase activity from Rhodobacter sphaeroides and its expression in Escherichia coli. J Bacteriol. 1987 Aug;169(8):3669–3678. doi: 10.1128/jb.169.8.3669-3678.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hallenbeck P. L., Lerchen R., Hessler P., Kaplan S. Phosphoribulokinase activity and regulation of CO2 fixation critical for photosynthetic growth of Rhodobacter sphaeroides. J Bacteriol. 1990 Apr;172(4):1749–1761. doi: 10.1128/jb.172.4.1749-1761.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hallenbeck P. L., Lerchen R., Hessler P., Kaplan S. Roles of CfxA, CfxB, and external electron acceptors in regulation of ribulose 1,5-bisphosphate carboxylase/oxygenase expression in Rhodobacter sphaeroides. J Bacteriol. 1990 Apr;172(4):1736–1748. doi: 10.1128/jb.172.4.1736-1748.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Henikoff S., Haughn G. W., Calvo J. M., Wallace J. C. A large family of bacterial activator proteins. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6602–6606. doi: 10.1073/pnas.85.18.6602. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Henikoff S. Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene. 1984 Jun;28(3):351–359. doi: 10.1016/0378-1119(84)90153-7. [DOI] [PubMed] [Google Scholar]
  22. Joint I. R., Morris I., Fuller R. C. Purification of a complex of alkaline fructose 1,6-bisphosphatase and phosphoribulokinase from Rhodospirillum rubrum. J Biol Chem. 1972 Aug 10;247(15):4833–4838. [PubMed] [Google Scholar]
  23. Jouanneau Y., Tabita F. R. In vivo regulation of form I ribulose 1,5-bisphosphate carboxylase/oxygenase from Rhodopseudomonas sphaeroides. Arch Biochem Biophys. 1987 Apr;254(1):290–303. doi: 10.1016/0003-9861(87)90105-6. [DOI] [PubMed] [Google Scholar]
  24. Jouanneau Y., Tabita F. R. Independent regulation of synthesis of form I and form II ribulose bisphosphate carboxylase-oxygenase in Rhodopseudomonas sphaeroides. J Bacteriol. 1986 Feb;165(2):620–624. doi: 10.1128/jb.165.2.620-624.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kay R., McPherson J. Hybrid pUC vectors for addition of new restriction enzyme sites to the ends of DNA fragments. Nucleic Acids Res. 1987 Mar 25;15(6):2778–2778. doi: 10.1093/nar/15.6.2778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Keen N. T., Tamaki S., Kobayashi D., Trollinger D. Improved broad-host-range plasmids for DNA cloning in gram-negative bacteria. Gene. 1988 Oct 15;70(1):191–197. doi: 10.1016/0378-1119(88)90117-5. [DOI] [PubMed] [Google Scholar]
  27. Kusian B., Yoo J. G., Bednarski R., Bowien B. The Calvin cycle enzyme pentose-5-phosphate 3-epimerase is encoded within the cfx operons of the chemoautotroph Alcaligenes eutrophus. J Bacteriol. 1992 Nov;174(22):7337–7344. doi: 10.1128/jb.174.22.7337-7344.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. LASCELLES J. The formation of ribulose 1:5-diphosphate carboxylase by growing cultures of Athiorhodaceae. J Gen Microbiol. 1960 Dec;23:499–510. doi: 10.1099/00221287-23-3-499. [DOI] [PubMed] [Google Scholar]
  29. Leustek T., Hartwig R., Weissbach H., Brot N. Regulation of ribulose bisphosphate carboxylase expression in Rhodospirillum rubrum: characteristics of mRNA synthesized in vivo and in vitro. J Bacteriol. 1988 Sep;170(9):4065–4071. doi: 10.1128/jb.170.9.4065-4071.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Markwell M. A., Haas S. M., Bieber L. L., Tolbert N. E. A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal Biochem. 1978 Jun 15;87(1):206–210. doi: 10.1016/0003-2697(78)90586-9. [DOI] [PubMed] [Google Scholar]
  31. Meijer W. G., Arnberg A. C., Enequist H. G., Terpstra P., Lidstrom M. E., Dijkhuizen L. Identification and organization of carbon dioxide fixation genes in Xanthobacter flavus H4-14. Mol Gen Genet. 1991 Feb;225(2):320–330. doi: 10.1007/BF00269865. [DOI] [PubMed] [Google Scholar]
  32. ORMEROD J. G., ORMEROD K. S., GEST H. Light-dependent utilization of organic compounds and photoproduction of molecular hydrogen by photosynthetic bacteria; relationships with nitrogen metabolism. Arch Biochem Biophys. 1961 Sep;94:449–463. doi: 10.1016/0003-9861(61)90073-x. [DOI] [PubMed] [Google Scholar]
  33. Pridmore R. D. New and versatile cloning vectors with kanamycin-resistance marker. Gene. 1987;56(2-3):309–312. doi: 10.1016/0378-1119(87)90149-1. [DOI] [PubMed] [Google Scholar]
  34. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sarles L. S., Tabita F. R. Derepression of the synthesis of D-ribulose 1,5-bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum. J Bacteriol. 1983 Jan;153(1):458–464. doi: 10.1128/jb.153.1.458-464.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Schlaman H. R., Okker R. J., Lugtenberg B. J. Regulation of nodulation gene expression by NodD in rhizobia. J Bacteriol. 1992 Aug;174(16):5177–5182. doi: 10.1128/jb.174.16.5177-5182.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Schultz J. E., Weaver P. F. Fermentation and anaerobic respiration by Rhodospirillum rubrum and Rhodopseudomonas capsulata. J Bacteriol. 1982 Jan;149(1):181–190. doi: 10.1128/jb.149.1.181-190.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Stanier R. Y., Doudoroff M., Kunisawa R., Contopoulou R. THE ROLE OF ORGANIC SUBSTRATES IN BACTERIAL PHOTOSYNTHESIS. Proc Natl Acad Sci U S A. 1959 Aug;45(8):1246–1260. doi: 10.1073/pnas.45.8.1246. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Tabita F. R., Gibson J. L., Bowien B., Dijkhuizen L., Meijer W. G. Uniform designation for genes of the Calvin-Benson-Bassham reductive pentose phosphate pathway of bacteria. FEMS Microbiol Lett. 1992 Dec 1;78(2-3):107–110. doi: 10.1111/j.1574-6968.1992.tb05551.x. [DOI] [PubMed] [Google Scholar]
  40. Tabita F. R., McFadden B. A. D-ribulose 1,5-diphosphate carboxylase from Rhodospirillum rubrum. II. Quaternary structure, composition, catalytic, and immunological properties. J Biol Chem. 1974 Jun 10;249(11):3459–3464. [PubMed] [Google Scholar]
  41. Taylor L. A., Rose R. E. A correction in the nucleotide sequence of the Tn903 kanamycin resistance determinant in pUC4K. Nucleic Acids Res. 1988 Jan 11;16(1):358–358. doi: 10.1093/nar/16.1.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Viale A. M., Kobayashi H., Akazawa T., Henikoff S. rbcR [correction of rcbR], a gene coding for a member of the LysR family of transcriptional regulators, is located upstream of the expressed set of ribulose 1,5-bisphosphate carboxylase/oxygenase genes in the photosynthetic bacterium Chromatium vinosum. J Bacteriol. 1991 Aug;173(16):5224–5229. doi: 10.1128/jb.173.16.5224-5229.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  44. Wang X., Falcone D. L., Tabita F. R. Reductive pentose phosphate-independent CO2 fixation in Rhodobacter sphaeroides and evidence that ribulose bisphosphate carboxylase/oxygenase activity serves to maintain the redox balance of the cell. J Bacteriol. 1993 Jun;175(11):3372–3379. doi: 10.1128/jb.175.11.3372-3379.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Weaver K. E., Tabita F. R. Isolation and partial characterization of Rhodopseudomonas sphaeroides mutants defective in the regulation of ribulose bisphosphate carboxylase/oxygenase. J Bacteriol. 1983 Nov;156(2):507–515. doi: 10.1128/jb.156.2.507-515.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Whitman W., Tabita F. R. Inhibition of D-ribulose 1,5-bisphosphate carboxylase by pyridoxal 5'-phosphate. Biochem Biophys Res Commun. 1976 Aug 23;71(4):1034–1039. doi: 10.1016/0006-291x(76)90758-0. [DOI] [PubMed] [Google Scholar]
  47. Windhövel U., Bowien B. Identification of cfxR, an activator gene of autotrophic CO2 fixation in Alcaligenes eutrophus. Mol Microbiol. 1991 Nov;5(11):2695–2705. doi: 10.1111/j.1365-2958.1991.tb01978.x. [DOI] [PubMed] [Google Scholar]
  48. Windhövel U., Bowien B. On the operon structure of the cfx gene clusters in Alcaligenes eutrophus. Arch Microbiol. 1990;154(1):85–91. doi: 10.1007/BF00249183. [DOI] [PubMed] [Google Scholar]
  49. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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