Mutant Protease Acidic Stability via Position 303 Substitution

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Solution Overview

Problem

Existing liquid detergents face challenges in stabilizing proteases at acidic pH due to the presence of surfactants and other components, which affects their washing performance.

Innovation Solution

A mutant protease with an amino acid sequence having at least 90% identity to a specific sequence (SEQ ID NO:2) is developed, where the amino acid residue at position 303 is substituted with specific amino acids to enhance stability under acidic conditions.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If a protease with alkaline optimum pH is used in liquid detergent, then washing performance on composite stains is improved, but stability in acidic liquid detergent is worsened

Engineering Contradiction:
Improvewashing performanceVSAvoidenzyme stability
Core Design Contradiction:
ReliabilityVSStability of the object's composition

Solution Approach 1:

The invention changes the pH stability parameter of the protease by introducing specific amino acid substitutions (G303D, G303E, G303F, G303H, G303I, G303M, G303N, G303P, G303Q, G303T, G303V, G303W, or G303Y). These substitutions modify the enzyme's structural properties to expand its pH stability range from alkaline only to both acidic and alkaline conditions, while maintaining high protease activity across a broader pH spectrum including pH 3-11.

Inventive Principle:
Principle #35Parameter changes

2Adaptability or versatility

If liquid detergent contains surfactants and other components, then various detergent functions are achieved, but enzyme preservation stability is worsened

Engineering Contradiction:
Improvedetergent functionVSAvoidenzyme preservation
Core Design Contradiction:
Adaptability or versatilityVSReliability

Solution Approach 1:

The invention modifies the protease's structural parameters through amino acid substitutions at position 303, enhancing its resistance to denaturation by surfactants and other detergent components. The mutant proteases demonstrate improved stability in the presence of cationic surfactants and other liquid detergent ingredients, maintaining enzymatic activity where wild-type proteases would be inactivated.

Inventive Principle:
Principle #35Parameter changes

3Object-generated harmful factors

If pH of liquid detergent is made weakly acidic or strongly acidic to reduce smell, then antibacterial property is improved, but protease stability is worsened

Engineering Contradiction:
Improvesmell reductionVSAvoidprotease stability
Core Design Contradiction:
Object-generated harmful factorsVSStability of the object's composition

Solution Approach 1:

The invention expands the pH stability window of the protease through specific amino acid substitutions, enabling the enzyme to remain stable and active in weakly acidic to strongly acidic conditions (pH 3-6). This allows liquid detergents to be formulated at acidic pH levels for smell reduction and enhanced antibacterial properties while maintaining effective protease activity for stain removal.

Inventive Principle:
Principle #35Parameter changes

Data Source

PatentUS12319943B2Mutant protease
Publication Date: 2025.06.03 KAO CORP

AI summary

Provided is a mutant protease with improved stability under acidic condition. A mutant protease that consists of the amino acid sequence as shown in SEQ ID NO:2 or an amino acid sequence having an identity of at least 90% thereto, in which the amino acid residue at a position corresponding to position 303 in the amino acid sequence as shown in SEQ ID NO:2 is substituted with another amino acid residue.