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. 1997 Aug;179(16):5131–5137. doi: 10.1128/jb.179.16.5131-5137.1997

Cloning and characterization of the gene (farA) encoding the receptor for an extracellular regulatory factor (IM-2) from Streptomyces sp. strain FRI-5.

M Waki 1, T Nihira 1, Y Yamada 1
PMCID: PMC179372  PMID: 9260956

Abstract

IM-2 is a butyrolactone autoregulator that controls production of blue pigment and nucleoside antibiotics in Streptomyces sp. strain FRI-5. An IM-2-specific receptor gene, farA, was cloned from strain FRI-5, and nucleotide sequencing revealed that the farA gene consists of 666 bp encoding a 221-amino-acid protein of 24.3 kDa with an NH2-terminal amino acid sequence identical to that of purified native receptor. Another gene, farX, encoding a homolog of AfsA of Streptomyces griseus, was present upstream of farA. The monocistronic nature of the farA transcript was shown by Northern blot hybridization, and the transcript level increased upon addition of IM-2. Recombinant FarA expressed in and purified from E. coli showed clear ligand specificity toward IM-2, with a dissociation constant (Kd) for [3H]IM-2-C5 of 18.2 nM. FarA showed high overall homology to BarA (virginiae butanolide receptor from S. virginiae) and ArpA (A-factor receptor from S. griseus). Sequence alignment of the three receptor proteins revealed that the NH2-terminal region containing a helix-turn-helix DNA binding motif was highly conserved. The DNA binding motif is common in procaryotic repressors of the TetR family, suggesting that all the Streptomyces autoregulator receptors may act as transcriptional repressors.

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Selected References

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  1. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  2. Grant S. G., Jessee J., Bloom F. R., Hanahan D. Differential plasmid rescue from transgenic mouse DNAs into Escherichia coli methylation-restriction mutants. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4645–4649. doi: 10.1073/pnas.87.12.4645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gräfe U., Schade W., Eritt I., Fleck W. F., Radics L. A new inducer of anthracycline biosynthesis from Streptomyces viridochromogenes. J Antibiot (Tokyo) 1982 Dec;35(12):1722–1723. doi: 10.7164/antibiotics.35.1722. [DOI] [PubMed] [Google Scholar]
  4. Hopwood D. A., Kieser T., Wright H. M., Bibb M. J. Plasmids, recombination and chromosome mapping in Streptomyces lividans 66. J Gen Microbiol. 1983 Jul;129(7):2257–2269. doi: 10.1099/00221287-129-7-2257. [DOI] [PubMed] [Google Scholar]
  5. Horinouchi S., Suzuki H., Nishiyama M., Beppu T. Nucleotide sequence and transcriptional analysis of the Streptomyces griseus gene (afsA) responsible for A-factor biosynthesis. J Bacteriol. 1989 Feb;171(2):1206–1210. doi: 10.1128/jb.171.2.1206-1210.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Kim H. S., Nihira T., Tada H., Yanagimoto M., Yamada Y. Identification of binding protein of virginiae butanolide C, an autoregulator in virginiamycin production, from Streptomyces virginiae. J Antibiot (Tokyo) 1989 May;42(5):769–778. doi: 10.7164/antibiotics.42.769. [DOI] [PubMed] [Google Scholar]
  7. Kim H. S., Tada H., Nihira T., Yamada Y. Purification and characterization of virginiae butanolide C-binding protein, a possible pleiotropic signal-transducer in Streptomyces virginiae. J Antibiot (Tokyo) 1990 Jun;43(6):692–706. doi: 10.7164/antibiotics.43.692. [DOI] [PubMed] [Google Scholar]
  8. Kondo K., Higuchi Y., Sakuda S., Nihira T., Yamada Y. New virginiae butanolides from Streptomyces virginiae. J Antibiot (Tokyo) 1989 Dec;42(12):1873–1876. doi: 10.7164/antibiotics.42.1873. [DOI] [PubMed] [Google Scholar]
  9. Lipman D. J., Pearson W. R. Rapid and sensitive protein similarity searches. Science. 1985 Mar 22;227(4693):1435–1441. doi: 10.1126/science.2983426. [DOI] [PubMed] [Google Scholar]
  10. Miyake K., Horinouchi S., Yoshida M., Chiba N., Mori K., Nogawa N., Morikawa N., Beppu T. Detection and properties of A-factor-binding protein from Streptomyces griseus. J Bacteriol. 1989 Aug;171(8):4298–4302. doi: 10.1128/jb.171.8.4298-4302.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Nihira T., Shimizu Y., Kim H. S., Yamada Y. Structure-activity relationships of virginiae butanolide C, an inducer of virginiamycin production in Streptomyces virginiae. J Antibiot (Tokyo) 1988 Dec;41(12):1828–1837. doi: 10.7164/antibiotics.41.1828. [DOI] [PubMed] [Google Scholar]
  12. Okamoto S., Nakamura K., Nihira T., Yamada Y. Virginiae butanolide binding protein from Streptomyces virginiae. Evidence that VbrA is not the virginiae butanolide binding protein and reidentification of the true binding protein. J Biol Chem. 1995 May 19;270(20):12319–12326. doi: 10.1074/jbc.270.20.12319. [DOI] [PubMed] [Google Scholar]
  13. Onaka H., Ando N., Nihira T., Yamada Y., Beppu T., Horinouchi S. Cloning and characterization of the A-factor receptor gene from Streptomyces griseus. J Bacteriol. 1995 Nov;177(21):6083–6092. doi: 10.1128/jb.177.21.6083-6092.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Rao R. N., Richardson M. A., Kuhstoss S. Cosmid shuttle vectors for cloning and analysis of Streptomyces DNA. Methods Enzymol. 1987;153:166–198. doi: 10.1016/0076-6879(87)53053-1. [DOI] [PubMed] [Google Scholar]
  15. Ruengjitchatchawalya M., Nihira T., Yamada Y. Purification and characterization of the IM-2-binding protein from Streptomyces sp. strain FRI-5. J Bacteriol. 1995 Feb;177(3):551–557. doi: 10.1128/jb.177.3.551-557.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. 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]
  17. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  18. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  19. Ward J. M., Janssen G. R., Kieser T., Bibb M. J., Buttner M. J., Bibb M. J. Construction and characterisation of a series of multi-copy promoter-probe plasmid vectors for Streptomyces using the aminoglycoside phosphotransferase gene from Tn5 as indicator. Mol Gen Genet. 1986 Jun;203(3):468–478. doi: 10.1007/BF00422072. [DOI] [PubMed] [Google Scholar]
  20. Yamada Y., Sugamura K., Kondo K., Yanagimoto M., Okada H. The structure of inducing factors for virginiamycin production in Streptomyces virginiae. J Antibiot (Tokyo) 1987 Apr;40(4):496–504. doi: 10.7164/antibiotics.40.496. [DOI] [PubMed] [Google Scholar]

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