Chemistry: molecular biology and microbiology – Enzyme – proenzyme; compositions thereof; process for... – Hydrolase
Patent
1996-02-14
1999-01-05
Weber, Jon P.
Chemistry: molecular biology and microbiology
Enzyme , proenzyme; compositions thereof; process for...
Hydrolase
435220, 510300, 510305, 510306, C12N 954, C12N 952, C11O 3386
Patent
active
058561676
DESCRIPTION:
BRIEF SUMMARY
CROSS-REFERENCE TO RELATED APPLICATIONS
This application is a 371 of PCT/DK94/00331 filed Sep. 2, 1994, which is incorporated herein by reference.
FIELD OF INVENTION
This invention is in the field of proteases derived from strains of Bacillus sp. More specifically, the invention is directed towards a novel protease derived from a strain of a novel Bacillus sp., which is characterized by being stable in solutions containing hypochlorite and/or other oxidizing agents. Moreover, the invention is directed towards a process for the preparation of the protease, and the use of the protease in processes in which water containing hypochlorite is being used.
BACKGROUND OF THE INVENTION
Proteases have been marketed for more than 20 years for a lot of different purposes, the most important as being ingredients in detergents.
Proteases have been developed by isolation of proteases found in nature. Most commercially available proteases are obtained from the genus Bacillus. Currently new types of proteases enter the market, offering the possibility of giving a better cost/performance ratio at various specified conditions.
Examples of commercial Bacillus protease products are Alcalase.RTM., Esperase.RTM., Primase.RTM., Savinase.RTM. and Durazyme.RTM. (a protein-engineered variant of Savinase), all available from Novo Nordisk A/S, Denmark. These and similar enzyme products from other commercial sources are active in detergent solutions, i.e. at pH values in the range from 8 to 11 and in the presence of sequestering agents, surfactants and bleaching agents such as sodium borate, but their activity decreases if the process water used contains hypochlorite. This is an increasing problem as more and more water in the industrialized world gets chlorinated.
SUMMARY OF THE INVENTION
It is an object of the present invention to provide novel proteases with improved stability performance in solutions containing hypochlorite.
Accordingly, in its first aspect, the invention provides a protease having immunochemical properties identical to those of a protease derived from the strain Bacillus sp., DSM 8473, the protease being stable in solutions containing hypochlorite.
In a second aspect, the invention relates to a biologically pure culture of a strain of a novel Bacillus sp. In a more specific aspect, the invention relates to a strain of Bacillus sp., DSM 8473, or a mutant or a variant thereof.
In a third aspect, the invention provides a process for the preparation of the protease, which process comprises cultivation of a protease producing strain of a novel Bacillus sp. in a suitable nutrient medium, containing carbon and nitrogen sources and inorganic salts, followed by recovery of the desired enzyme. In a more specific aspect, Bacillus sp., DSM 8473, or a mutant or a variant thereof encoding a protease having immunochemical properties identical to those of the protease derived from Bacillus sp., DSM 8473, is cultivated.
In a fourth aspect, the use of the protease in processes in which water containing hypochlorite is being used, is claimed:
BRIEF DESCRIPTION OF DRAWINGS
The present invention is further illustrated by reference to the accompanying drawings, in which
FIG. 1 shows the relation between temperature and the proteolytic activity of a novel protease according to the invention (the protease preparation obtained according to Ex.1, with 2% of casein as substrate and at pH 9.5).
FIG. 2 shows the relation between pH and the proteolytic activity of a novel protease according to the invention (the protease preparation obtained according to Ex.1, with 2% of casein as substrate and at 25.degree. C., using Britten-Robinson buffers adjusted to predetermined pH values in the pH range of from 6 to 11).
DETAILED DISCLOSURE OF THE INVENTION
The novel microorganism of the invention, able to produce an enzyme of the invention, is represented by the strain that was isolated from a soil sample.
The novel Bacillus sp. has been deposited according to the Budapest Treaty on the International Recognition of the Deposits of Microo
REFERENCES:
patent: 5118623 (1992-06-01), Boguslawski et al.
patent: 5358865 (1994-10-01), Outtrup et al.
Koltukova et al. (1985) Prikl. Biokhim. Microbiol., 21(4), "Some Physiochemical Properties of Bacillus mesentericus Proteinases", pp. 495-500, with English abstract.
Kalebina et al. (1983) Bioorg. Khim., 9(6), "Extracellular Serine Protease from Bacillus brevis -an Analog of Subtilisin BPN", pp. 815-823, with English abstract.
Chestukhina et al. (1982) Bioorg. Khim., 8(12), "Thiol Dependent Serine Proteinases 2: Extracellular Serine Proteinases", pp. 1649-1658, with English abstract.
Kalebina et al. (1988) Appl. Microbiol. Biotechnol., 28(6), "Serine Proteinase from Bacillus brevis : Lytic Action on Intact Yeast Cells", pp. 531-536.
Saravani et al. (1989) Biochem. Journal, 262(2), "Caldolase, A Chelator-Insensitive Extracellular Serine Proteinase from a Thermus spp. ", pp. 409-416.
Burton et al. (1993) Journal Gen. Microbiol., 139(6), "Purification and Characterization of a Serine Proteinase from Senescent Sporophores of the Commercial Mushroom Agaricus-bisporus", pp. 1379-1386.
Lambiris Elias J.
Novo Nordisk A S
Rozek Carol E.
Weber Jon P.
Zelson Steve T.
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