Skip to main content
SpringerLink
Log in

Novel stand-alone RAM domain protein-mediated catalytic control of anthranilate phosphoribosyltransferase in tryptophan biosynthesis in Thermus thermophilus

  • Special Feature: Original Paper
  • 11th International Congress on Extremophiles
  • Published:
Extremophiles Aims and scope Submit manuscript

Abstract

Regulation of amino acid metabolism (RAM) domains are widely distributed among prokaryotes. In most cases, a RAM domain fuses with a DNA-binding domain to act as a transcriptional regulator. The extremely thermophilic bacterium, Thermus thermophilus, only carries a single gene encoding a RAM domain-containing protein on its genome. This protein is a stand-alone RAM domain protein (SraA) lacking a DNA-binding domain. Therefore, we hypothesized that SraA, which senses amino acids through its RAM domain, may interact with other proteins to modify its functions. In the present study, we identified anthranilate phosphoribosyltransferase (AnPRT), the second enzyme in the tryptophan biosynthetic pathway, as a partner protein that interacted with SraA in T. thermophilus. In the presence of tryptophan, SraA was assembled to a decamer and exhibited the ability to form a stable hetero-complex with AnPRT. An enzyme assay revealed that AnPRT was only inhibited by tryptophan in the presence of SraA. This result suggests a novel feedback control mechanism for tryptophan biosynthesis through an inter-RAM domain interaction in bacteria.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

Abbreviations

PCR:

Polymerase chain reaction

SDS-PAGE:

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis

qRT-PCR:

Quantitative reverse transcriptase polymerase chain reaction

RAM domain:

Regulation of amino acid metabolism domain

Lrp:

Leucine-responsive regulatory protein

AS:

Anthranilate synthase

AnPRT:

Anthranilate phosphoribosyltransferase

PRPP:

Phosphoribosylpyrophosphate

References

  • Bae YM, Crawford IP (1990) The Rhizobium meliloti trpE(G) gene is regulated by attenuation, and its product, anthranilate synthase, is regulated by feedback inhibition. J Bacteriol 172:3318–3327

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Belser WL, Murphy JB, Delmer DP, Mills SE (1971) End product control of tryptophan biosynthesis in extracts and intact cells of the higher plant Nicotiana tabacum var. Wisconsin 38. Biochim Biophys Acta 237:1–10

    Article  CAS  PubMed  Google Scholar 

  • Brinkman AB, Bell SD, Lebbink RJ, de Vos WM, van der Oost J (2002) The Sulfolobus solfataricus Lrp-like protein LysM regulates lysine biosynthesis in response to lysine availability. J Biol Chem 277:29537–29549

    Article  CAS  PubMed  Google Scholar 

  • Brinkman AB, Ettema TJ, de Vos WM, van der Oost J (2003) The Lrp family of transcriptional regulators. Mol Microbiol 48:287–294

    Article  CAS  PubMed  Google Scholar 

  • Caligiuri MG, Bauerle R (1991) Identification of amino acid residues involved in feedback regulation of the anthranilate synthase complex from Salmonella typhimurium. Evidence for an amino-terminal regulatory site. J Biol Chem 266:8328–8335

    CAS  PubMed  Google Scholar 

  • Cava F, Hidalgo A, Berenguer J (2009) Thermus thermophilus as biological model. Extremophiles 13:213–231

    Article  CAS  PubMed  Google Scholar 

  • Chipman DM, Shaanan B (2001) The ACT domain family. Curr Opin Struct Biol 11:694–700

    Article  CAS  PubMed  Google Scholar 

  • Enoru-Eta J, Gigot D, Thia-Toong TL, Glansdorff N, Charlier D (2000) Purification and characterization of Sa-lrp, a DNA-binding protein from the extreme thermoacidophilic archaeon Sulfolobus acidocaldarius homologous to the bacterial global transcriptional regulator Lrp. J Bacteriol 182:3661–3672

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ernsting BR, Atkinson MR, Ninfa AJ, Matthews RG (1992) Characterization of the regulon controlled by the leucine-responsive regulatory protein in Escherichia coli. J Bacteriol 174:1109–1118

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ettema TJ, Brinkman AB, Tani TH, Rafferty JB, Van Der Oost J (2002) A novel ligand-binding domain involved in regulation of amino acid metabolism in prokaryotes. J Biol Chem 277:37464–37468

    Article  CAS  PubMed  Google Scholar 

  • Fujita A, Misumi Y, Honda S, Sato T, Koyama Y (2013) Construction of new cloning vectors that employ the phytoene synthase encoding gene for color screening of cloned DNA inserts in Thermus thermophilus. Gene 527:655–662

    Article  CAS  PubMed  Google Scholar 

  • Grant GA (2006) The ACT domain: a small molecule binding domain and its role as a common regulatory element. J Biol Chem 281:33825–33829

    Article  CAS  PubMed  Google Scholar 

  • Grove TH, Levy HR (1975) Fluorescent assay of anthranilate synthetase-anthranilate 5-phosphoribosylpyrophosphate phosphoribosyltransferase enzyme-complex on polyacrylamide gels. Anal Biochem 65:458–465

    Article  CAS  PubMed  Google Scholar 

  • Heery DM, Dunican LK (1993) Cloning of the trp gene cluster from a tryptophan-hyperproducing strain of Corynebacterium glutamicum: identification of a mutation in the trp leader sequence. Appl Environ Microbiol 59:791–799

    CAS  PubMed  PubMed Central  Google Scholar 

  • Henderson EJ, Nagano H, Zalkin H, Hwang LH (1970) The anthranilate synthetase-anthranilate 5-phosphoribosylpyrophosphate phosphoribosyltransferase aggregate. Purification of the aggregate and regulatory properties of anthranilate synthetase. J Biol Chem 245:1416–1423

    CAS  PubMed  Google Scholar 

  • Henne A, Bruggemann H, Raasch C et al (2004) The genome sequence of the extreme thermophile Thermus thermophilus. Nat Biotechnol 22:547–553

    Article  CAS  PubMed  Google Scholar 

  • Hove-Jensen B (1989) Phosphoribosylpyrophosphate (PRPP)-less mutants of Escherichia coli. Mol Microbiol 3:1487–1492

    Article  CAS  PubMed  Google Scholar 

  • Jackson EN, Yanofsky C (1974) Localization of two functions of the phosphoribosyl anthranilate transferase of Escherichia coli to distinct regions of the polypeptide chain. J Bacteriol 117:502–508

    CAS  PubMed  PubMed Central  Google Scholar 

  • Knoten CA, Hudson LL, Coleman JP, Farrow JM, Pesci EC (2011) KynR, a Lrp/AsnC-type transcriptional regulator, directly controls the kynurenine pathway in Pseudomonas aeruginosa. J Bacteriol 193:6567–6575

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Koyama Y, Hoshino T, Tomizuka N, Furukawa K (1986) Genetic-transformation of the extreme thermophile Thermus thermophilus and of other Thermus spp. J Bacteriol 166:338–340

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Matsui K, Miwa K, Sano K (1987) Two single-base-pair substitutions causing desensitization to tryptophan feedback inhibition of anthranilate synthase and enhanced expression of tryptophan genes of Brevibacterium lactofermentum. J Bacteriol 169:5330–5332

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Morollo AA, Eck MJ (2001) Structure of the cooperative allosteric anthranilate synthase from Salmonella typhimurium. Nat Struct Biol 8:243–247

    Article  CAS  PubMed  Google Scholar 

  • Nakamura A, Takakura Y, Kobayashi H, Hoshino T (2005) In vivo directed evolution for thermostabilization of Escherichia coli hygromycin B phosphotransferase and the use of the gene as a selection marker in the host-vector system of Thermus thermophilus. J Biosci Bioeng 100:158–163

    Article  CAS  PubMed  Google Scholar 

  • Nakano N, Okazaki N, Satoh S, Takio K, Kuramitsu S, Shinkai A, Yokoyama S (2006) Structure of the stand-alone RAM-domain protein from Thermus thermophilus HB8. Acta Crystallogr F 62:855–860

    Article  CAS  Google Scholar 

  • O’Gara JP, Dunican LK (1995) Mutations in the trpD gene of Corynebacterium glutamicum confer 5-methyltryptophan resistance by encoding a feedback-resistant anthranilate phosphoribosyltransferase. Appl Environ Microbiol 61:4477–4479

    PubMed  PubMed Central  Google Scholar 

  • Okamura H, Yokoyama K, Koike H, Yamada M, Shimowasa A, Kabasawa M, Kawashima T, Suzuki M (2007) A structural code for discriminating between transcription signals revealed by the feast/famine regulatory protein DM1 in complex with ligands. Structure 15:1325–1338

    Article  CAS  PubMed  Google Scholar 

  • Peeters E, Charlier D (2010) The Lrp family of transcription regulators in archaea. Archaea 2010:750457

    Article  PubMed  PubMed Central  Google Scholar 

  • Sambrook JF, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor New York

    Google Scholar 

  • Song N, Cui Y, Li Z, Chen L, Liu S (2016) New Targets and cofactors for the transcription factor LrpA from Mycobacterium tuberculosis. DNA Cell Biol 35:167–176

    Article  CAS  PubMed  Google Scholar 

  • Sugimoto S, Shiio I (1983) Regulation of tryptophan biosynthesis by feedback inhibition of the 2nd-step enzyme, anthranilate phosphoribosyl-transferase, in Brevibacterium flavum. Agric Biol Chem 47:2295–2305

    CAS  Google Scholar 

  • Tanaka T, Kawano N, Oshima T (1981) Cloning of 3-isopropylmalate dehydrogenase gene of an extreme thermophile and partial-purification of the gene-product. J Biochem (Tokyo) 89:677–682

    CAS  Google Scholar 

  • Tutino ML, Tosco A, Marino G, Sannia G (1997) Expression of Sulfolobus solfataricus trpE and trpG genes in E. coli. Biochem Biophys Res Commun 230:306–310

    Article  CAS  PubMed  Google Scholar 

  • Yokoyama K, Ishijima SA, Koike H, Kurihara C, Shimowasa A, Kabasawa M, Kawashima T, Suzuki M (2007) Feast/famine regulation by transcription factor FL11 for the survival of the hyperthermophilic archaeon Pyrococcus OT3. Structure 15:1542–1554

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported in part by JSPS KAKENHI Grant Nos. 24580137 (T.T.) and 15K07382 (T.T.). We thank Kaori Ohtsuki, Masaya Usui, and Aya Abe at the RRC center of RIKEN-BSI for their technical assistance with the MS analysis.

Author information

Author notes

    Authors and Affiliations

    1. Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo, 113-8657, Japan

      Tetsuo Kubota, Hajime Matsushita, Takeo Tomita, Saori Kosono, Tomohisa Kuzuyama & Makoto Nishiyama

    2. RIKEN Center for Sustainable Resource Science, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan

      Minoru Yoshida

    Authors
    1. Tetsuo Kubota
      View author publications

      You can also search for this author in PubMed Google Scholar

    2. Hajime Matsushita
      View author publications

      You can also search for this author in PubMed Google Scholar

    3. Takeo Tomita
      View author publications

      You can also search for this author in PubMed Google Scholar

    4. Saori Kosono
      View author publications

      You can also search for this author in PubMed Google Scholar

    5. Minoru Yoshida
      View author publications

      You can also search for this author in PubMed Google Scholar

    6. Tomohisa Kuzuyama
      View author publications

      You can also search for this author in PubMed Google Scholar

    7. Makoto Nishiyama
      View author publications

      You can also search for this author in PubMed Google Scholar

    Corresponding author

    Correspondence to Makoto Nishiyama.

    Additional information

    Communicated by H. Atomi.

    T. Kubota and H. Matsushita contributed equally to this work.

    This article is part of a special feature based on the 11th International Congress on Extremophiles held in Kyoto, Japan, September 12–16, 2016.

    Electronic supplementary material

    Below is the link to the electronic supplementary material.

    Supplementary material 1 (PDF 325 kb)

    Rights and permissions

    Reprints and permissions

    About this article

    Check for updates. Verify currency and authenticity via CrossMark

    Cite this article

    Kubota, T., Matsushita, H., Tomita, T. et al. Novel stand-alone RAM domain protein-mediated catalytic control of anthranilate phosphoribosyltransferase in tryptophan biosynthesis in Thermus thermophilus . Extremophiles 21, 73–83 (2017). https://doi.org/10.1007/s00792-016-0884-0

    Download citation

    • Received:

    • Accepted:

    • Published:

    • Issue Date:

    • DOI: https://doi.org/10.1007/s00792-016-0884-0

    Keywords

    Search

    Navigation