Subtilase Variants

a technology of subtilase and variants, applied in the field of new products, can solve the problems of not exactly known which physical or chemical characteristics, and not giving data in respect of such variants, so as to improve the performance of detergents and improve storage stability

Inactive Publication Date: 2009-06-25
NOVOZYMES AS
View PDF49 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0069]It has now surprisingly been found that a subtilase variant having improved storage stability and/or improved performance in detergents, can be obtained by substituting one or more amino acid residues situated in, or in the vicinity of a hydrophobic domain of the parent subtilase for an amino acid residue more hydrophobic than the original residue, sa...

Problems solved by technology

Also, it is still not exactly known which physical or chemical characteristics are responsible for a good washing performance or stability of a protease in a specific detergent composition.
Since these positions all except position −1 were known to be involved in the functioning of the ...

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Subtilase Variants
  • Subtilase Variants
  • Subtilase Variants

Examples

Experimental program
Comparison scheme
Effect test

example 1

Construction and Expression of Enzyme Variants

[0316]A vector suited to a synthetic gene coding for subtilase 309 and its mutants was constructed. It is essentially a pUC19 plasmid [Yanish-Perron and Messing (1985) Gene; 33 103-119], in which the multiple cloning site has been replaced by a linker containing the restriction sites used to separate five sub-fragments constituting the gene. The new linker was inserted into EcoRI-HindIII cut pUC19 thereby destroying these sites. The details of this construction are described in WO 92 / 19729 on pages 25-26 and in FIG. 1 (sheets 1 / 7-7 / 7) thereof, the content of which is hereby included by reference.

[0317]Each subfragment was made from 6 to 12 oligonucleotides. The oligonucleotides were synthesized on an automatic DNA synthesizer using phosphoramidite chemistry on a controlled glass support [Beaucage and Carruthers (1981); Tetrahedron Letters 22 1859-1869].

[0318]The five subfragments were isolated on a 2% agarose gel and inserted into pSX191...

example 2

Purification of Enzyme Variants

[0324]This procedure relates to purification of a 10 liter scale fermentation of subtilisin 147, subtilisin 309 or mutants thereof.

[0325]Approximately 8 liters of fermentation broth were centrifuged at 5000 rpm for 35 minutes in 1 liter beakers. The supernatants were adjusted to pH 6.5 using 10% acetic acid and filtered on Seitz Supra S100 filter plates.

[0326]The filtrates were concentrated to approximately 400 ml using an Amicon CH2A UF unit equipped with an Amicon S1Y10 UF cartridge. The UF concentrate was centrifuged and filtered prior to absorption at room temperature on a Bacitracin affinity column at pH 7. The protease was eluted from the Bacitracin column at room temperature using 25% 2-propanol and 1 M sodium chloride in a buffer solution with 0.01 dimethylglutaric acid, 0.1 M boric acid and 0.002 M calcium chloride adjusted to pH 7.

[0327]The fractions with protease activity from the Bacitracin purification step were combined and applied to a 7...

example 3

Stability in Detergent Compositions Comprising Enzyme Variants

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The present invention relates to enzymes produced by mutating the genes for a number of subtilases and expressing the mutated genes in suitable hosts are presented. The enzymes exhibit improved stability and/or improved wash performance in any detergent in comparison to their wild type parent enzymes. The enzymes are well-suited for use in any detergent and for some in especially liquid or solid shaped detergent compositions.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is a continuation of application Ser. No. 10 / 618,934 filed Jul. 14, 2003, which is a continuation of application Ser. No. 09 / 587,747 filed Jun. 5, 2000, now U.S. Pat. No. 6,682,924, which is a continuation of application Ser. No. 09 / 120,577 filed Jul. 22, 1998, now U.S. Pat. No. 6,190,900, which is a continuation of application Ser. No. 08 / 642,987 filed May 6, 1996, now U.S. Pat. No. 5,837,517, which claims priority under 35 U.S.C. 119 of Danish application nos. 0519 / 95 and 0421 / 96 filed May 5, 1995 and Apr. 12, 1996, respectively, and European application no. 95201161 filed May 5, 1995, the contents of which are fully incorporated herein by reference.REFERENCE TO A SEQUENCE LISTING[0002]This application contains a paper copy and computer readable form of a Sequence Listing, which are incorporated herein by reference.BACKGROUND OF THE INVENTION[0003]1. Field of the Invention[0004]This invention relates to novel mutant enz...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C11D3/386C12N9/54C12N9/56C12S11/00C12N1/21C12N15/57
CPCC12N9/54C11D3/386
Inventor SIERKSTRA, LAURENS NICOLAASKLUGKIST, JANMARKVARDSEN, PETERVON DER OSTEN, CLAUSBAUDITZ, PETER
Owner NOVOZYMES AS
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap