Characterization of Two Cel (Cellulose Degradation) Genes of Clostridium Thermocellum Coding for Endoglucanases

Abstract

Two structural genes, celA and celB, coding respectively for the endoglucanases A and B of Clostridium thermocellum, were subcloned from cosmids of a gene bank into plasmid pBR322. Both genes are carried in a DNA sequence of about 2.7 kb. They share no homology and are not contiguous on the C. thermocellum chromosome. They are expressed in E. coli independently of their orientation in the cloning vector.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1

    Ng, T.K., Weimer, P.J. and Zeikus, J.G. 1977. Cellulolytic and physiological properties of Cloitridium thermocellum. Arch. Microbiol. 114: 1–7.

  2. 2

    Zeikus, J.G. 1980. Chemical and fuel production by anaerobic bacteria. Annu. Rev. Microbiol. 34: 423–464.

  3. 3

    Aït, N., Creuzet, N. and Forget, P. 1979. Partial purification of cellulase from Clostridium thermocellum. J. Gen. Microbiol. 113: 399–402.

  4. 4

    Shinmyo, A., Garcia-Mardnez, D.V. and Demain, A.L. 1979. Studies on the extracellular cellulolytic enzyme complex produced by Clostridium thermocellum. 1979. J. Appl. Biochem. 1: 202–209.

  5. 5

    Garcia-Martinez, D.V., Shinmyo, A., Madia, A. and Demain, A.L. 1980. Studies on the cellulase produced by Clostridium thermocellum. European J. Appl. Microbiol. Biotechnol. 9: 189–197.

  6. 6

    Petre, J., Longin, R. and Millet, J. 1981. Purification and properties of an endo-β-1,4-glucanase from Clostridium thermocellum. Biochimie 63: 629–639.

  7. 7

    Ng, T.K. and Zeikus, J.G. 1981. Purification and characterization of an endoglucanase (1,4-β-D-glucan glucanohydrolase) from Clostridium thermocellum. Biochem. J. 199: 341–350.

  8. 8

    Creuzet, N. and Frixon, C. 1983. Purification and characterization of an endoglucanase from a newly isolated thermophilic anaerobic bacterium. Biochimie 65: 149–156.

  9. 9

    Aït, N., Creuzet, N. and Cattaneo, J. 1982. Properties of β-glucosidase purified from Clostridium thermocellum. J. Gen. Microbiol. 128: 569–577.

  10. 10

    Cornet, P., Tronik, D., Millet, J. and Aubert, J.-P. 1983. Cloning and expression in Escherichia coli of Clostridium thermocellum genes coding for amino acid synthesis and cellulose hydrolysis. FEMS Microbiol. Letters 16: 137–141.

  11. 11

    Teather, R.M. and Wood, P.J. 1982. Use of Congo red-polysaccharide interactions in enumeration and characterization of cellulolytic bacteria from the bovine rumen. Appl. Environ. Microbiol. 43: 777–780.

  12. 12

    Uhlin, B.E. and Nordstrom, K. 1977. R plasmid gene dosage effects in Escherichia coli K-12: copy mutants of the R plasmid RIdrd-19. Plasmid 1: 1–7.

  13. 13

    Whittle, D.J., Kilburn, D.G., Warren, R.A.J. and Miller, R.C. Jr., 1982. Molecular cloning of a Cellulomonas fimi cellulase gene in Escherichia coli. Gene 17: 139–145.

  14. 14

    Collmer, A. and Wilson, D.B. 1983. Cloning and Expression of a Thermonumospora YX Endocellulase Gene in E. coli. BIO/TECHNOLOGY 1: 594–601.

  15. 15

    Boyer, H.W. and Roulland-Dussoix, D. 1969. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J. Mol. Biol. 41: 459–472.

  16. 16

    Reeve, J.N. 1977. Bacteriophage infection of minicells. A general method for identification of in vivo bacteriophage directed polypeptide biosynthesis. Mol. Gen. Genet. 158: 73–79.

  17. 17

    Bolivar, F., Rodriguez, R.L., Greene, J.P., Betlach, M.C., Heyneker, H.L. and Boyer, H.W. 1977. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene 2: 95–113.

  18. 18

    Miller, J.H. 1972. In Experiments in Molecular Genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York.

  19. 19

    Humphreys, G.O., Willshaw, G.A. and Anderson, E.S. 1975. A simple method for the preparation of large quantities of pure plasmid DNA. Biochim. Biophys. Acta 382: 457–463.

  20. 20

    Birnboim, H.C. and Doly, J. 1979. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 7: 1513–1523.

  21. 21

    Maniatis, T., Fritsch, E.F. and Sambrook, J. 1982. Molecular Cloning, a Laboratory Manual. Cold Spring Harbor Laboratory, New York.

  22. 22

    Cohen, S.N., Chang, A.C.Y. and Hsu, L. 1972. Non chromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA. Proc. Natl. Acad. Sci. (USA) 69: 2110–2114.

  23. 23

    Rigby, P.W.J., Dieckmann, M., Rhodes, C. and Berg, P. 1977. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J. Mol. Biol. 113: 237–251.

  24. 24

    Southern, E. 1975. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. Mol. Biol. 98: 503–517.

  25. 25

    Quiviger, B., Franche, C., Lutfalla, G., Rice, D., Haselkorn, R. and Elmerich, C. 1982. Cloning of a nitrogen-fixation (nif) gene cluster of Azaspirillum brasilense. Biochimie 64: 495–502.

  26. 26

    Kessler, S.W. 1976. Cell membrane antigen isolation with the staphylococcal protein A-antibody adsorbent. J. Immunol. 117: 1482–1496.

  27. 27

    Béguin, P., Nicolas, P., Boussetta, H., Fahy, C. and Cohen, P. 1981. Characterization of the 80,000 molecular weight form of neurophysin isolated from bovine neurohypophysis. J. Biol. Chem. 256: 9289–9294.

  28. 28

    Pardee, A.B., Jacob, F. and Monod, J. 1959. The genetic control and cytoplasmic expression of “inducibility” in the synthesis of β-galactosidase by Escherichia coli. J. Mol. Biol. 1: 165–178.

  29. 29

    Ross, G.W. and O'Callaghan, C.H. 1975. β-lactamase assays. In Methods in Enzymology, J. H. Hash (ed.), Academic Press, New York 43: 69–85.

  30. 30

    Heppel, L.A. 1967. Selective release of enzymes from bacteria. Science 156: 1451–1455.

  31. 31

    Sedmak, J.J. and Grossberg, S.E. 1977. A rapid, sensitive and versatile assay for protein using Coomassie brilliant blue G250. Anal. Biochem. 79: 544–552.

  32. 32

    Lammli, U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.

Download references

Author information

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Cornet, P., Millet, J., Béguin, P. et al. Characterization of Two Cel (Cellulose Degradation) Genes of Clostridium Thermocellum Coding for Endoglucanases. Nat Biotechnol 1, 589–594 (1983). https://doi.org/10.1038/nbt0983-589

Download citation

Further reading