Abstract
From a hot spring of the Azores islands a novel thermophilic bacterium belonging to the Thermotogales order was isolated. This strain, which grows optimally at 70(deg)C and pH 6.5, is the first known extreme thermophile that is able to degrade native feathers at high temperatures. The enzyme system converts feather meal to amino acids and peptides. On the basis of physiological, morphological, and 16S rDNA studies the new isolate was found to be a member of the Thermotogales order and was identified as Fervidobacterium pennavorans. The strain was highly related to Fervidobacterium islandicum and Fervidobacterium pullulanolyticum. The cell-bound keratinolytic enzyme system was purified 32-fold by detergent treatment with CHAPS (3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propanesulfonate) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was characterized as a serine protease with a molecular mass of 130 kDa and an isoelectric point of 3.8. Optimal activity was measured at 80(deg)C and pH 10.0. Furthermore, 19 anaerobic thermophilic archaea and bacteria belonging to the orders Thermococcales, Thermoproteales, Thermotogales, and Clostridiales (growth temperatures between 60 and 105(deg)C) were tested for their abilities to grow on feathers and produce heat-stable keratinolytic enzymes. None of the tested extremophilic microorganisms was able to attack the substrate in a native form.
Full Text
The Full Text of this article is available as a PDF (488.0 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Asahi M., Lindquist R., Fukuyama K., Apodaca G., Epstein W. L., McKerrow J. H. Purification and characterization of major extracellular proteinases from Trichophyton rubrum. Biochem J. 1985 Nov 15;232(1):139–144. doi: 10.1042/bj2320139. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Balch W. E., Fox G. E., Magrum L. J., Woese C. R., Wolfe R. S. Methanogens: reevaluation of a unique biological group. Microbiol Rev. 1979 Jun;43(2):260–296. doi: 10.1128/mr.43.2.260-296.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Behn W., Arnold C. G. Die Wirkung von Streptomycin and Neamin auf die Chloroplasten- und Mitochondrienstruktur von Chlamydomonas reinhardii. Protoplasma. 1974;82(1):77–89. doi: 10.1007/BF01276872. [DOI] [PubMed] [Google Scholar]
- 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]
- Böckle B., Galunsky B., Müller R. Characterization of a keratinolytic serine proteinase from Streptomyces pactum DSM 40530. Appl Environ Microbiol. 1995 Oct;61(10):3705–3710. doi: 10.1128/aem.61.10.3705-3710.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cashion P., Holder-Franklin M. A., McCully J., Franklin M. A rapid method for the base ratio determination of bacterial DNA. Anal Biochem. 1977 Aug;81(2):461–466. doi: 10.1016/0003-2697(77)90720-5. [DOI] [PubMed] [Google Scholar]
- 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]
- Ebeling W., Hennrich N., Klockow M., Metz H., Orth H. D., Lang H. Proteinase K from Tritirachium album Limber. Eur J Biochem. 1974 Aug 15;47(1):91–97. doi: 10.1111/j.1432-1033.1974.tb03671.x. [DOI] [PubMed] [Google Scholar]
- Heussen C., Dowdle E. B. Electrophoretic analysis of plasminogen activators in polyacrylamide gels containing sodium dodecyl sulfate and copolymerized substrates. Anal Biochem. 1980 Feb;102(1):196–202. doi: 10.1016/0003-2697(80)90338-3. [DOI] [PubMed] [Google Scholar]
- Kitadokoro K., Tsuzuki H., Nakamura E., Sato T., Teraoka H. Purification, characterization, primary structure, crystallization and preliminary crystallographic study of a serine proteinase from Streptomyces fradiae ATCC 14544. Eur J Biochem. 1994 Feb 15;220(1):55–61. doi: 10.1111/j.1432-1033.1994.tb18598.x. [DOI] [PubMed] [Google Scholar]
- Klingeberg M., Galunsky B., Sjoholm C., Kasche V., Antranikian G. Purification and Properties of a Highly Thermostable, Sodium Dodecyl Sulfate-Resistant and Stereospecific Proteinase from the Extremely Thermophilic Archaeon Thermococcus stetteri. Appl Environ Microbiol. 1995 Aug;61(8):3098–3104. doi: 10.1128/aem.61.8.3098-3104.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kunert J. Keratin decomposition by dermatophytes. I. Sulfite production as a possible way of substrate denaturation. Z Allg Mikrobiol. 1973;13(6):489–498. doi: 10.1002/jobm.3630130606. [DOI] [PubMed] [Google Scholar]
- Kunert J. Keratin decomposition by dermatophytes: evidence of the sulphitolysis of the protein. Experientia. 1972 Sep 15;28(9):1025–1026. doi: 10.1007/BF01918649. [DOI] [PubMed] [Google Scholar]
- Kunert J., Krajcí D. An electron microscopy study of keratin degradation by the fungus Microsporum gypseum in vitro. Mykosen. 1981 Aug;24(8):485–496. doi: 10.1111/j.1439-0507.1981.tb01898.x. [DOI] [PubMed] [Google Scholar]
- 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]
- Lin X., Kelemen D. W., Miller E. S., Shih J. C. Nucleotide sequence and expression of kerA, the gene encoding a keratinolytic protease of Bacillus licheniformis PWD-1. Appl Environ Microbiol. 1995 Apr;61(4):1469–1474. doi: 10.1128/aem.61.4.1469-1474.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lin X., Lee C. G., Casale E. S., Shih J. C. Purification and Characterization of a Keratinase from a Feather-Degrading Bacillus licheniformis Strain. Appl Environ Microbiol. 1992 Oct;58(10):3271–3275. doi: 10.1128/aem.58.10.3271-3275.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lin X., Tang J. Purification, characterization, and gene cloning of thermopsin, a thermostable acid protease from Sulfolobus acidocaldarius. J Biol Chem. 1990 Jan 25;265(3):1490–1495. [PubMed] [Google Scholar]
- Mukhopadhyay R. P., Chandra A. L. Keratinase of a streptomycete. Indian J Exp Biol. 1990 Jun;28(6):575–577. [PubMed] [Google Scholar]
- REYNOLDS E. S. The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol. 1963 Apr;17:208–212. doi: 10.1083/jcb.17.1.208. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rainey F. A., Fritze D., Stackebrandt E. The phylogenetic diversity of thermophilic members of the genus Bacillus as revealed by 16S rDNA analysis. FEMS Microbiol Lett. 1994 Jan 15;115(2-3):205–211. doi: 10.1111/j.1574-6968.1994.tb06639.x. [DOI] [PubMed] [Google Scholar]
- Schumann J., Wrba A., Jaenicke R., Stetter K. O. Topographical and enzymatic characterization of amylases from the extremely thermophilic eubacterium Thermotoga maritima. FEBS Lett. 1991 Apr 22;282(1):122–126. doi: 10.1016/0014-5793(91)80459-g. [DOI] [PubMed] [Google Scholar]
- Sinha U., Wolz S. A., Lad P. J. Two new extracellular serine proteases from Streptomyces fradiae. Int J Biochem. 1991;23(10):979–984. doi: 10.1016/0020-711x(91)90133-8. [DOI] [PubMed] [Google Scholar]
- Spurr A. R. A low-viscosity epoxy resin embedding medium for electron microscopy. J Ultrastruct Res. 1969 Jan;26(1):31–43. doi: 10.1016/s0022-5320(69)90033-1. [DOI] [PubMed] [Google Scholar]
- Takami H., Akiba T., Horikoshi K. Characterization of an alkaline protease from Bacillus sp. no. AH-101. Appl Microbiol Biotechnol. 1990 Aug;33(5):519–523. doi: 10.1007/BF00172544. [DOI] [PubMed] [Google Scholar]
- VENABLE J. H., COGGESHALL R. A SIMPLIFIED LEAD CITRATE STAIN FOR USE IN ELECTRON MICROSCOPY. J Cell Biol. 1965 May;25:407–408. doi: 10.1083/jcb.25.2.407. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Voss T., Melchers K., Scheirle G., Schäfer K. P. Structural comparison of recombinant pulmonary surfactant protein SP-A derived from two human coding sequences: implications for the chain composition of natural human SP-A. Am J Respir Cell Mol Biol. 1991 Jan;4(1):88–94. doi: 10.1165/ajrcmb/4.1.88. [DOI] [PubMed] [Google Scholar]
- Wawrzkiewicz K., Lobarzewski J., Wolski T. Intracellular keratinase of Trichophyton gallinae. J Med Vet Mycol. 1987 Aug;25(4):261–268. [PubMed] [Google Scholar]
- Williams C. M., Richter C. S., Mackenzie J. M., Shih J. C. Isolation, identification, and characterization of a feather-degrading bacterium. Appl Environ Microbiol. 1990 Jun;56(6):1509–1515. doi: 10.1128/aem.56.6.1509-1515.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Winterhalter C., Liebl W. Two Extremely Thermostable Xylanases of the Hyperthermophilic Bacterium Thermotoga maritima MSB8. Appl Environ Microbiol. 1995 May;61(5):1810–1815. doi: 10.1128/aem.61.5.1810-1815.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wolin E. A., Wolfe R. S., Wolin M. J. Viologen dye inhibition of methane formation by Methanobacillus omelianskii. J Bacteriol. 1964 May;87(5):993–998. doi: 10.1128/jb.87.5.993-998.1964. [DOI] [PMC free article] [PubMed] [Google Scholar]