Bacillus pumilus protease amino acid substitutions for liquid detergent stability
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Solution Overview
Problem
Existing proteases used in washing and cleaning agents often lack sufficient catalytic activity and storage stability, particularly in liquid surfactant-containing preparations, leading to suboptimal cleaning performance on protein-sensitive soils under standard washing conditions.
Innovation Solution
A protease from Bacillus pumilus with specific amino acid substitutions, such as 9T, 144K, 252T, and 271E, and additional substitutions at various positions, enhancing its cleaning performance and storage stability, is developed.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If wild-type protease from Bacillus pumilus is used in liquid surfactant-containing preparations, then the formulation can be stored, but the protease lacks sufficient catalytic activity and storage stability under standard washing conditions
Solution Approach 1:
The patent applies parameter changes by introducing specific amino acid substitutions (9T, 144K, 252T, 271E and additional substitutions at positions 18, 38, 48, 89, 101, 131, 145, 147, 149, 162, 166, 172, 189, 192, 205, 211, 215, 217, 218, 224 and 274) into the protease sequence. These molecular parameter changes optimize the enzyme's structure and function, simultaneously improving both storage stability and catalytic activity in liquid surfactant-containing preparations under standard washing conditions.
2Productivity
If protease is optimized for high catalytic activity, then cleaning performance improves, but storage stability deteriorates
Solution Approach 1:
The patent resolves this contradiction through targeted parameter changes in the protease amino acid sequence. The specific substitutions (9T, 144K, 252T, 271E and others) are designed to simultaneously enhance catalytic activity and maintain storage stability, achieving both improvements without the usual trade-off between these two properties.
3Ease of operation
If protease is used in liquid formulations with surfactants, then the formulation is practical for washing, but the protease exhibits insufficient proteolytic activity under standard washing conditions
Solution Approach 1:
The patent addresses this contradiction by implementing specific amino acid substitutions in the protease sequence that enhance its proteolytic activity while maintaining compatibility with liquid surfactant-containing formulations. The modifications (9T, 144K, 252T, 271E and additional positions) ensure the enzyme remains active and effective under standard washing conditions, resolving the issue of insufficient proteolytic activity in practical formulations.
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 modified protease exhibits improved proteolytic activity and increased stability, achieving at least 101% of the reference protease's performance and maintaining high cleaning efficacy over extended storage periods, even at temperatures between 20°C to 40°C.
Implementation Method 1
They bring about the decomposition of protein-containing soilings on the item to be cleaned. Of these, in turn, proteases of the subtilisin type (subtilases, subtilopeptidases, EC 3.4.21.62) are particularly important and are serine proteases due to the catalytically active amino acids. They act as non-specific endopeptidases and hydrolyze any acid amide bonds that are inside peptides or proteins.
Implementation Method 2
They act as non-specific endopeptidases and hydrolyze any acid amide bonds that are inside peptides or proteins.
Data Source
AI summary
The invention relates to proteases comprising an amino acid sequence which has at least 70% sequence identity with the amino acid sequence given in SEQ ID NO:1 over its entire length and has, in each case based on the numbering according to SEQ ID NO:1, (i) amino acid substitutions, preferably selected from the amino acid substitutions 9T, 144K, 252T and 271E, at the positions corresponding to positions 9, 144, 252 and 271; and (ii) at least one further amino acid substitution at at least one of the positions corresponding to positions 18, 38, 48, 89, 101, 131, 145, 147, 149, 162, 166, 172, 189, 192, 205, 211, 215, 217, 218, 224 and 274. The invention also relates to the production and use of said proteases. Proteases of this type exhibit a very good cleaning performance.