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Metalloprotease from chryseobacterium

一种蛋白酶活性、选自的技术，应用在水解酶、洗涤剂混合物组合物的制备、生物化学设备和方法等方向，能够解决金属蛋白酶不稳定等问题

Pending Publication Date: 2016-11-23
NOVOZYMES AS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] However, known metalloproteases are very unstable under normal wash conditions and in conventional detergent compositions
Therefore, the use of metalloproteases in detergents and in washing and cleaning processes has not been commercialized

Method used

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  • Metalloprotease from chryseobacterium
  • Metalloprotease from chryseobacterium
  • Metalloprotease from chryseobacterium

Examples

Experimental program
Comparison scheme
Effect test

example

[0346] Materials and methods

[0347] Materials used for washing assays

[0348] Grass stain sample 062KC was obtained from Warwick Equest Ltd, Unit 55, Consett Business Park, Consett, County Durham, DH8 6BN England.

[0349] Grass Stain Samples NZ grass was prepared by using juice from whole grass mixed on WFK20A cotton / polyester.

[0350] The remaining samples were obtained from the Center For Testmaterials BV, PO Box 120, 3133 KT Vlaardingen, The Netherlands.

[0351]

[0352] standard detergent

[0353] Use standard detergents B and J. Standard Detergent B is similar to a European type liquid detergent composition. Standard Detergent J is similar to US type liquid detergent compositions.

[0354]

[0355]

[0356] washing assay

[0357] Automated Mechanical Stress Assay (AMSA) for Laundry

[0358] To assess wash performance, laundry washing experiments were performed using the Automated Mechanical Stress Assay (AMSA). With AMSA, the wash performance of...

example 1

[0392] Example 1. Cloning and expression of the M4 metalloprotease from Chryseobacterium sp. 10696

[0393] This metalloprotease was derived from a bacterial strain received as NCIMB1314; it was originally isolated from a sample of minced fish meat and identified as Flavobacterium. Chromosomal DNA from pure cultures was purified and subjected to whole genome sequencing using Illumina technology. The assembled genome sequence and subsequent analysis of the 16S ribosomal subunit gene sequence indicated that the correct taxonomic identification was as a new species in the genus Chryseobacterium and the strain was named Chryseobacterium 10696.

[0394] Genomic sequence analysis of metalloproteases from the MEROPS family M4 was performed by comparison with the M4 protease NprE from Bacillus subtilis 168 (Uniprot P68736) by searching using the BLAST program. This analysis identified that the gene encoding the putative M4 metalloprotease has the nucleotide sequence given in SEQ ID:1...

example 2

[0400] Example 2. Purification of M4 protease from Chryseobacterium sp. 10696

[0401] The supernatant was filtered through a Nalgene 0.2 μm filter unit to remove remaining Bacillus host cells. The solid (NH 4 ) 2 SO 4 Add to 0.2μm filtrate to reach 1.5M (NH 4 ) 2 SO 4 final concentration, and the M4 protease solution was applied to 100mM H 3 BO 3 , 10mM MES, 2mM CaCl 2 , 1.5M (NH 4 ) 2 SO 4 , on a Phenyl-Toyopearl column equilibrated in pH 6 (from TosoHaas). After washing the column well with equilibration buffer, the M4 protease was washed with 100 mM H 3 BO 3 , 10mM MES, 2mM CaCl 2 , pH 6 for elution. The elution peak was applied to 100mM H 3 BO 3 , 10mM MES, 2mM CaCl 2 , on a subtilisin sepharose column (from Upfront chromatography) equilibrated in pH 6. After washing the column well with equilibration buffer, the M4 protease was washed with 100 mM H with 25% (v / v) 2-propanol. 3 BO 3 , 10mM MES, 2mM CaCl 2 , 1M NaCl, pH 6 elution. The eluted peaks we...

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Abstract

The present invention relates to a new metalloprotease and the use thereof in cleaning processes, such as laundry and dish wash. The invention also relates to detergent compositions and cleaning compositions comprising a metalloprotease.

Description

[0001] References to Sequence Listings [0002] This application contains a Sequence Listing in computer readable form. This computer readable form is hereby incorporated by reference. field of invention [0003] The present invention relates to cleaning and / or detergent compositions comprising a metalloprotease (E.C 3.4.24). The present invention further relates to novel metalloproteases from the genus Chryseobacterium and their use in cleaning processes such as dishwashing and laundry washing. In addition, the present invention relates to methods of performing cleaning such as dishwashing and laundry. Background of the invention [0004] For more than 30 years, the detergent industry has employed different enzymes in detergent formulations, the most commonly used enzymes include proteases, amylases and lipases, each suitable for removing different types of soils. Detergent compositions typically include a complex combination of ingredients in addition to these enzymes. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/52C12N15/57C11D3/386
CPCC11D3/386C12N9/52C11D2111/12
Inventor M.杰曼森S.尼曼德-格拉鲁普P.R.奥斯特加德A.V.赖泽J.比耶尔
Owner NOVOZYMES AS
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