Engineered Polypeptides for Low-Lactose GOS Production
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current enzymes used for producing galacto-oligosaccharides, such as those from Bifidobacterium bifidum, require high lactose concentrations or excess acceptor molecules, limiting their efficiency in dairy products like milk, where lactose levels are low.
Innovation Solution
Development of polypeptides with a high ratio of transgalactosylation to hydrolysis activity, specifically designed for Bacillus subtilis expression, which can efficiently convert lactose into galacto-oligosaccharides even at low lactose levels, enhancing the growth of beneficial microbial strains like Bifidobacterium.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If current enzymes from Bifidobacterium bifidum are used for producing GOS, then transgalactosylation activity is achieved, but high lactose concentrations or excess acceptor molecules are required, limiting efficiency in dairy products
Solution Approach 1:
The patent modifies the enzyme's amino acid sequence through specific mutations (e.g., E365Q, E366Q substitutions) to change its catalytic properties. These parameter changes in the enzyme structure enable it to function effectively at lower lactose concentrations and with lower acceptor molecule requirements, directly resolving the contradiction between productivity and substance quantity requirements
Solution Approach 2:
The patent creates engineered variants that copy the essential transgalactosylation function of the original Bifidobacterium bifidum enzyme while improving its performance characteristics. By copying and optimizing the catalytic core with specific mutations, the enzyme achieves higher efficiency at lower substrate concentrations, suitable for dairy product applications
2Productivity
If high lactose concentrations are used for GOS production, then transgalactosylation activity increases, but the applicability to dairy products like milk is limited due to low natural lactose levels
Solution Approach 1:
The enzyme engineering modifies catalytic parameters through amino acid substitutions (e.g., E365Q, E366Q) to shift the enzyme's optimal operating conditions. This enables the enzyme to maintain high transgalactosylation activity at the low lactose concentrations naturally present in dairy products, thereby improving both productivity and adaptability simultaneously
3Productivity
If excess acceptor molecules are used to produce GOS, then transgalactosylation efficiency improves, but production cost and process complexity increase
Solution Approach 1:
The patent engineers enzyme variants with modified amino acid sequences (e.g., E365Q, E366Q mutations) that change the enzyme's substrate binding characteristics. These parameter changes enable the enzyme to achieve high transgalactosylation efficiency with minimal or no excess acceptor molecules, simplifying the manufacturing process while maintaining productivity
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
These polypeptides effectively produce galacto-oligosaccharides in situ in dairy products, reducing free lactose levels and promoting the growth of health-promoting bacteria, offering a cheaper and more efficient method for producing these beneficial compounds.
Implementation Method 1
the enzyme hydrolyses lactose and transiently binds the galactose monosaccharide in a galactose-enzyme complex that transfers galactose to the hydroxyl group of water, resulting in the liberation of D-galactose and D-glucose. However, at high lactose concentrations some β-galactosidases are able to transfer galactose to the hydroxyl groups of D-galactose or D-glucose in a process called transgalactosylation whereby galacto-oligosaccharides are produced
Implementation Method 2
The enzyme β-galactosidase (EC 3.2.1.23) usually hydrolyses lactose to the monosaccharides D-glucose and D-galactose
Data Source
AI summary
The present invention relates to polypeptides, specifically polypeptides having transgalactosylating activity and nucleic acids encoding these, and their uses in e.g. dairy product.