Novel Pseudomonas Stutzeri Heparinases for Selective Lysis
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Solution Overview
Problem
Current heparinases, such as those from Flavobacterium heparinum, have limitations in selectivity and specificity for lysing heparin, necessitating the discovery of new enzymes with improved properties for quality testing and production of low molecular weight heparin.
Innovation Solution
A novel Pseudomonas stutzeri strain (Z7) is isolated and used to produce two intracellular heparinases, PShepI and PShepII, with distinct physicochemical properties and high selectivity for heparin lysing, which are purified using methods involving ammonium sulfate precipitation, anion exchange, cation exchange, and affinity chromatography.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If conventional heparinases from Flavobacterium heparinum are used, then heparin can be lysed to produce low molecular weight heparin, but the selectivity and specificity for lysing heparin is insufficient
Solution Approach 1:
The patent discovers and characterizes novel heparinases with different physicochemical parameters (molecular weight, isoelectric point, optimal pH, optimal temperature) compared to conventional heparinases. These parameter changes result in improved selectivity and specificity for lysing heparin while maintaining high catalytic efficiency. The new enzymes exhibit distinct kinetic properties including different Km and Vmax values, enabling optimized performance for specific applications.
2Reliability
If new heparinases are discovered to improve selectivity, then the quality testing and production capabilities are enhanced, but the complexity of enzyme selection and characterization increases
Solution Approach 1:
The patent identifies and characterizes multiple distinct heparinase enzymes (heparinase I, II, and III) from Flavobacterium heparinum, each with unique properties. This segmentation allows selection of the most appropriate enzyme for specific quality testing or production applications, enhancing reliability while managing complexity through systematic classification and characterization of each enzyme variant.
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 novel heparinases exhibit enhanced selectivity and efficiency in lysing heparin, enabling effective quality testing and production of low or ultra-low molecular weight heparin, with specific characteristics like molecular weight, isoelectric point, and substrate activity ratios.
Implementation Method 1
Heparinase refers to an enzyme that is able to specifically break the glucosidic bond on the backbone of heparin and heparinoid
Implementation Method 2
The lysis can be carried out by a variety of methods known in the art for disrupting cells, such as freeze-thaw lysis, sonication, high pressure treatment, and the like
Data Source
AI summary
The invention relates to the field of bioengineering. In particular, the invention relates to a heparinase-producing Pseudomonas stutzeri strain and a heparinase derived therefrom. Furthermore, the invention relates to the preparation and use of the heparinase.