Skip to main content
Biochemical Journal logoLink to Biochemical Journal
. 1991 Aug 1;277(Pt 3):887–890. doi: 10.1042/bj2770887

Thermostable cellobiohydrolase from the thermophilic eubacterium Thermotoga sp. strain FjSS3-B.1. Purification and properties.

L D Ruttersmith 1, R M Daniel 1
PMCID: PMC1151328  PMID: 1872819

Abstract

Exo-1,4-beta-cellobiohydrolase (EC 3.2.1.91) was isolated from the culture supernatant of Thermotoga sp. strain FjSS3-B.1, an extremely thermophilic eubacterium that grows optimally at 80 degrees C. The enzyme was purified to homogeneity as determined by SDS/PAGE and has an Mr of 36,000. The enzyme is the most thermostable cellulase reported to date, with a half-life at 108 degrees C of 70 min in buffer. In a 40 min assay, maximal activity was recorded at 105 degrees C. Cellobiohydrolase from strain FjSS3-B.1 is active against amorphous cellulose and CM-cellulose but only effects limited hydrolysis of filter paper or Sigmacell 20. The only product identified by h.p.l.c. is the disaccharide cellobiose. The enzyme has a pH optimum around neutral and is stabilized by the presence of 0.8 M-NaCl.

Full text

PDF
887

Selected References

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

  1. Ahern T. J., Klibanov A. M. The mechanisms of irreversible enzyme inactivation at 100C. Science. 1985 Jun 14;228(4705):1280–1284. doi: 10.1126/science.4001942. [DOI] [PubMed] [Google Scholar]
  2. Ayala G., de Gómez-Puyou M. T., Gómez-Puyou A., Darszon A. Thermostability of membrane systems in organic solvents. FEBS Lett. 1986 Jul 14;203(1):41–43. doi: 10.1016/0014-5793(86)81432-6. [DOI] [PubMed] [Google Scholar]
  3. Blumentals I. I., Robinson A. S., Kelly R. M. Characterization of sodium dodecyl sulfate-resistant proteolytic activity in the hyperthermophilic archaebacterium Pyrococcus furiosus. Appl Environ Microbiol. 1990 Jul;56(7):1992–1998. doi: 10.1128/aem.56.7.1992-1998.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Brown S. H., Costantino H. R., Kelly R. M. Characterization of Amylolytic Enzyme Activities Associated with the Hyperthermophilic Archaebacterium Pyrococcus furiosus. Appl Environ Microbiol. 1990 Jul;56(7):1985–1991. doi: 10.1128/aem.56.7.1985-1991.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bryant F. O., Adams M. W. Characterization of hydrogenase from the hyperthermophilic archaebacterium, Pyrococcus furiosus. J Biol Chem. 1989 Mar 25;264(9):5070–5079. [PubMed] [Google Scholar]
  7. Cowan D. A., Smolenski K. A., Daniel R. M., Morgan H. W. An extremely thermostable extracellular proteinase from a strain of the archaebacterium Desulfurococcus growing at 88 degrees C. Biochem J. 1987 Oct 1;247(1):121–133. doi: 10.1042/bj2470121. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Eisenthal R., Cornish-Bowden A. The direct linear plot. A new graphical procedure for estimating enzyme kinetic parameters. Biochem J. 1974 Jun;139(3):715–720. doi: 10.1042/bj1390715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  10. Lever M. Colorimetric and fluorometric carbohydrate determination with p-hydroxybenzoic acid hydrazide. Biochem Med. 1973 Apr;7(2):274–281. doi: 10.1016/0006-2944(73)90083-5. [DOI] [PubMed] [Google Scholar]
  11. Ng T. K., Weimer T. K., Zeikus J. G. Cellulolytic and physiological properties of Clostridium thermocellum. Arch Microbiol. 1977 Jul 26;114(1):1–7. doi: 10.1007/BF00429622. [DOI] [PubMed] [Google Scholar]
  12. Reynolds P. H., Sissons C. H., Daniel R. M., Morgan H. W. Comparison of Cellulolytic Activities in Clostridium thermocellum and Three Thermophilic, Cellulolytic Anaerobes. Appl Environ Microbiol. 1986 Jan;51(1):12–17. doi: 10.1128/aem.51.1.12-17.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Rouvinen J., Bergfors T., Teeri T., Knowles J. K., Jones T. A. Three-dimensional structure of cellobiohydrolase II from Trichoderma reesei. Science. 1990 Jul 27;249(4967):380–386. doi: 10.1126/science.2377893. [DOI] [PubMed] [Google Scholar]
  14. Simpson H. D., Haufler U. R., Daniel R. M. An extremely thermostable xylanase from the thermophilic eubacterium Thermotoga. Biochem J. 1991 Jul 15;277(Pt 2):413–417. doi: 10.1042/bj2770413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Zaks A., Klibanov A. M. Enzymatic catalysis in organic media at 100 degrees C. Science. 1984 Jun 15;224(4654):1249–1251. doi: 10.1126/science.6729453. [DOI] [PubMed] [Google Scholar]
  16. Zale S. E., Klibanov A. M. Why does ribonuclease irreversibly inactivate at high temperatures? Biochemistry. 1986 Sep 23;25(19):5432–5444. doi: 10.1021/bi00367a014. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES