Skip to main content
NCBI home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1991 Aug;57(8):2121–2130. doi: 10.1128/aem.57.8.2121-2130.1991

Purification and Cooperative Activity of Enzymes Constituting the Xylan-Degrading System of Thermomonospora fusca

Susan L Bachmann 1, Alan J McCarthy 1,*
PMCID: PMC183538  PMID: 16348531

Abstract

The thermophilic actinomycete Thermomonospora fusca produced endoxylanase, α-arabinofuranosidase, β-xylosidase, and acetyl esterase activities maximally during growth on xylan. Growth yields on glucose, xylose, or arabinose were comparable, but production of endoxylanase and β-xylosidase was not induced on these substrates. The crude xylanase activity was thermostable and relatively resistant to end product inhibition by xylobiose and xylan hydrolysis products. Six proteins with xylanase activity were identified by zymogram analysis of isoelectric focusing gels, but only a 32-kDa protein exhibiting three isomeric forms could be purified by fast protein liquid chromatography. Endoglucanases were also identified in carboxymethylcellulose-grown cultures, and their distinction from endoxylanases was confirmed. α-Arabinofuranosidase activity was due to a single dimeric protein of 92 kDa, which was particularly resistant to end product inhibition by arabinose. Three bands of acetyl esterase activity were detected by zymogram analysis, and there was evidence that these mainly consisted of an intracellular 80-kDa protein secreted to yield active 40-kDa subunits in the culture supernatant. The acetyl esterases were found to be responsible for acetyl xylan esterase activity in T. fusca, in contrast to the distinction proposed in some other systems. The addition of purified βxylosidase to endoxylanase increased the hydrolysis of xylan, probably by relieving end product inhibition. The enhanced saccharification of wheat straw caused by the addition of purified α-arabinofuranosidase to T. fusca endoxylanase suggested a truly synergistic relationship, in agreement with proposals that arabinose side groups on the xylan chain participate in cross-linking within the plant cell wall structure.

Full text

PDF
2121

Images in this article

2124Image
on p.2124
2125Image
on p.2125
2126Image
on p.2126
2126Image
on p.2126

Selected References

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

  1. Fleming G., Dawson M. T., Patching J. W. The isolation of strains of Bacillus subtilis showing improved plasmid stability characteristics by means of selective chemostat culture. J Gen Microbiol. 1988 Aug;134(8):2095–2101. doi: 10.1099/00221287-134-8-2095. [DOI] [PubMed] [Google Scholar]
  2. Ghangas G. S., Hu Y. J., Wilson D. B. Cloning of a Thermomonospora fusca xylanase gene and its expression in Escherichia coli and Streptomyces lividans. J Bacteriol. 1989 Jun;171(6):2963–2969. doi: 10.1128/jb.171.6.2963-2969.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ghangas G. S., Wilson D. B. Cloning of the Thermomonospora fusca Endoglucanase E2 Gene in Streptomyces lividans: Affinity Purification and Functional Domains of the Cloned Gene Product. Appl Environ Microbiol. 1988 Oct;54(10):2521–2526. doi: 10.1128/aem.54.10.2521-2526.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Ghangas G. S., Wilson D. B. Expression of a Thermomonospora fusca Cellulase Gene in Streptomyces lividans and Bacillus subtilis. Appl Environ Microbiol. 1987 Jul;53(7):1470–1475. doi: 10.1128/aem.53.7.1470-1475.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Greve L. C., Labavitch J. M., Hungate R. E. alpha-L-arabinofuranosidase from Ruminococcus albus 8: purification and possible role in hydrolysis of alfalfa cell wall. Appl Environ Microbiol. 1984 May;47(5):1135–1140. doi: 10.1128/aem.47.5.1135-1140.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hägerdal B., Harris H., Pye E. K. Association of beta-glucosidase with intact cells of Thermoactinomyces. Biotechnol Bioeng. 1979 Mar;21(3):345–355. doi: 10.1002/bit.260210302. [DOI] [PubMed] [Google Scholar]
  7. Kaji A., Yoshihara O. Properties of purified -L-arabinofuranosidase from Corticium rolfsii. Biochim Biophys Acta. 1971 Nov 13;250(2):367–371. doi: 10.1016/0005-2744(71)90193-8. [DOI] [PubMed] [Google Scholar]
  8. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [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. Lee S. F., Forsberg C. W., Rattray J. B. Purification and Characterization of Two Endoxylanases from Clostridium acetobutylicum ATCC 824. Appl Environ Microbiol. 1987 Apr;53(4):644–650. doi: 10.1128/aem.53.4.644-650.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lim K. B., Doraisingham S., Thirumoorthy T., Lee C. T., Ling A. E., Tan T. Foscarnet (phosphonoformate sodium) in the treatment of recurrent male genital herpes. Ann Acad Med Singapore. 1986 Oct;15(4):617–622. [PubMed] [Google Scholar]
  12. Lowe S. E., Theodorou M. K., Trinci A. P. Cellulases and xylanase of an anaerobic rumen fungus grown on wheat straw, wheat straw holocellulose, cellulose, and xylan. Appl Environ Microbiol. 1987 Jun;53(6):1216–1223. doi: 10.1128/aem.53.6.1216-1223.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mackenzie C. R., Bilous D., Schneider H., Johnson K. G. Induction of Cellulolytic and Xylanolytic Enzyme Systems in Streptomyces spp. Appl Environ Microbiol. 1987 Dec;53(12):2835–2839. doi: 10.1128/aem.53.12.2835-2839.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. McDermid K. P., Mackenzie C. R., Forsberg C. W. Esterase Activities of Fibrobacter succinogenes subsp. succinogenes S85. Appl Environ Microbiol. 1990 Jan;56(1):127–132. doi: 10.1128/aem.56.1.127-132.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ramachandra M., Crawford D. L., Pometto A. L. Extracellular Enzyme Activities during Lignocellulose Degradation by Streptomyces spp.: A Comparative Study of Wild-Type and Genetically Manipulated Strains. Appl Environ Microbiol. 1987 Dec;53(12):2754–2760. doi: 10.1128/aem.53.12.2754-2760.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Reilly P. J. Xylanases: structure and function. Basic Life Sci. 1981;18:111–129. doi: 10.1007/978-1-4684-3980-9_8. [DOI] [PubMed] [Google Scholar]
  17. Teather R. M., Wood P. J. Use of Congo red-polysaccharide interactions in enumeration and characterization of cellulolytic bacteria from the bovine rumen. Appl Environ Microbiol. 1982 Apr;43(4):777–780. doi: 10.1128/aem.43.4.777-780.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Wong K. K., Tan L. U., Saddler J. N. Multiplicity of beta-1,4-xylanase in microorganisms: functions and applications. Microbiol Rev. 1988 Sep;52(3):305–317. doi: 10.1128/mr.52.3.305-317.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES