Thermolysin Variant Stability in Liquid Detergents

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Solution Overview

Problem

Current detergents face challenges in removing complex stains, maintaining fabric color and tensile strength, preventing dye transfer, and providing anti-wrinkle, antibobbling, and anti-shrinkage properties, while also dealing with residue build-up and discoloration due to the destabilizing effects of builders and metal chelators on proteases.

Innovation Solution

A thermolysin-like neutral protease enzyme variant with improved storage stability and catalytic activity, specifically a Geobacillus thermolysin variant with amino acid substitutions at position 151, is used in cleaning compositions, along with stabilizers like borax and zinc ions, and additional enzymes to enhance cleaning performance.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If builders and metal chelators are added to liquid detergents to enhance cleaning performance, then stain removal capability is improved, but protease stability and activity deteriorate due to destabilizing effects

Engineering Contradiction:
Improvestain removal capabilityVSAvoidprotease stability
Core Design Contradiction:
ProductivityVSReliability

Solution Approach 1:

The patent applies parameter changes by modifying the amino acid sequence of thermolysin through site-directed mutagenesis, specifically changing residues at positions 151, 152, and 153. This creates variant enzymes with altered biochemical properties that confer resistance to inhibition by metal chelators and builders, thereby maintaining protease stability while allowing the use of these cleaning enhancement agents in the detergent formulation.

Inventive Principle:
Principle #35Parameter changes

Solution Approach 2:

The patent creates copies of the thermolysin enzyme with modified amino acid sequences. By synthesizing variant proteins that replicate the core catalytic function but with altered stability properties, the invention enables the enzyme to withstand the presence of metal chelators and builders without losing activity, thus resolving the contradiction between cleaning performance and enzyme stability.

Inventive Principle:
Principle #26Copying

2Reliability

If amino acid substitutions are made at position 151 of thermolysin to improve stability, then storage stability is improved, but catalytic activity may be affected

Engineering Contradiction:
Improvestorage stabilityVSAvoidcatalytic activity
Core Design Contradiction:
ReliabilityVSProductivity

Solution Approach 1:

The patent systematically explores parameter changes by testing multiple amino acid substitutions at position 151 (including H, Y, F, W, L, I, V, M, A, G, S, T, N, D, E, Q, R, K, C, P). Through this parameter optimization approach, the invention identifies specific variants that achieve the desired balance between enhanced storage stability and maintained or improved catalytic activity, as measured by casein degradation assays.

Inventive Principle:
Principle #35Parameter changes

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

The improved thermolysin variant maintains enhanced stability and performance under various conditions, effectively removing stains, preserving fabric color and strength, and providing additional benefits like static control and fabric softness in detergents.

Implementation Method 1

thermolysin-like neutral protease enzyme... Geobacillus thermolysin variant... removing stains

Methodology Applied
Scientific EffectEnzyme catalysis: Enzyme

Implementation Method 2

neutral protease enzyme... catalytic activity... removing stains

Methodology Applied
Scientific EffectProteolysis: Hydrolysis

Implementation Method 3

stabilizers like borax and zinc ions... maintains enhanced stability

Methodology Applied
Scientific EffectMetal ion stabilization:

Data Source

PatentEP2845900B1Production of thermolysin and variants thereof, and use in liquid detergents
Publication Date: 2019.01.09 DANISCO US INC
  • EP2845900B1 patent drawingFigure 1~2
  • EP2845900B1 patent drawingFigure 3
  • EP2845900B1 patent drawingFigure 4A

AI summary

The present invention provides methods and compositions comprising at least one thermolysin-like neutral protease enzyme with improved storage stability and/or catalytic activity. In some embodiments, the thermolysin finds use in cleaning and other applications comprising detergent. In some particularly preferred embodiments, the present invention provides methods and compositions comprising thermolysin formulated and/or engineered to resist detergent-induced inactivation.