Thermostable Asparaginase Variants for Acrylamide Reduction

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Solution Overview

Problem

Current asparaginases lack improved thermotolerance, which is necessary for effectively reducing acrylamide formation in foods during heating, as they are not stable at high temperatures, limiting their efficacy in food processing.

Innovation Solution

Development of novel asparaginase variants with modified amino acid residues at specific positions, such as 54, 57, 70, 83, 84, 86, 102, 137, 164, 196, 201, 228, 260, 262, 278, 283, 290, 307, 312, 323, 327, 334, 336, 337, 349, 351, 353, 366, and 375, enhancing their thermostability and activity at high temperatures.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If conventional asparaginases are used to reduce asparagine in food materials, then acrylamide formation is reduced, but the enzymes lack stability at high temperatures during heating processes

Engineering Contradiction:
ImprovethermostabilityVSAvoidactivity at high temperature
Core Design Contradiction:
ReliabilityVSTemperature

Solution Approach 1:

The patent applies parameter changes by modifying amino acid residues at specific positions (54, 57, 70, 83, 84, 86, 102, 137, 164, 196, 201, 228, 260, 262, 278, 283, 290, 307, 312, 323, 327, 334, 336, 337, 349, 351, 353, 366, and 375) in the asparaginase sequence to enhance thermostability. These substitutions alter the physical and chemical properties of the enzyme, allowing it to maintain stability and activity at elevated temperatures during food processing while continuing to effectively reduce acrylamide formation.

Inventive Principle:
Principle #35Parameter changes

2Object-affected harmful factors

If the enzyme is added prior to heating step to reduce amino acids involved in acrylamide formation, then acrylamide levels are reduced, but the enzyme activity is limited by poor thermotolerance during the heating process

Engineering Contradiction:
Improveacrylamide formationVSAvoidenzyme stability during heating
Core Design Contradiction:
Object-affected harmful factorsVSReliability

Solution Approach 1:

The patent modifies the enzyme's amino acid sequence to change its thermal stability parameters, enabling it to withstand the heating conditions used in food processing. This allows the enzyme to be added prior to heating and remain stable and active throughout the thermal processing step, effectively reducing acrylamide formation without being degraded by the heat.

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 modified asparaginases demonstrate improved residual activity and increased activity at elevated temperatures, effectively reducing acrylamide formation in foods, making them suitable for use in food processing to minimize acrylamide levels in thermally processed products.

Implementation Method 1

asparaginases having improved properties, preferably improved thermotolerance, such as improved activity at high temperatures and/or improved thermostability

Methodology Applied
Scientific EffectEnzyme catalysis: Enzyme

Implementation Method 2

reduction of acrylamide in foods

Methodology Applied
Scientific EffectHydrolysis: Hydrolysis

Data Source

PatentEP2137307B1Thermostable asparaginases
Publication Date: 2014.07.30 NOVOZYMES AS
  • EP2137307B1 patent drawingFigure 1
  • EP2137307B1 patent drawingFigure 2
  • EP2137307B1 patent drawingFigure 3

AI summary

The invention relates to new asparaginases having improved properties, preferably improved thermotolerance, such as improved activity at high temperatures and/or improved thermostability. The invention also relates to DNA sequences encoding such improved asparaginases, their production in a recombinant host cell, as well as methods of using the asparaginases, in particular for reduction of acrylamide in foods. The invention furthermore relates to methods of generating and preparing asparaginase variants having improved properties.