Coagulation Factor X Activating Enzyme Purification for Scale-Up
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Solution Overview
Problem
Existing methods for isolating coagulation factor X activating enzyme from Russell's viper venom yield low purity and activity, making large-scale production challenging.
Innovation Solution
A novel coagulation factor X activating enzyme with specific glycosylation modifications and a purification method involving sequential anion and cation exchange chromatography, enhancing purity and activity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If traditional isolation methods (Sephadex G-150 size exclusion chromatography, QAE-Sephadex A-50 anion chromatography) are used to prepare coagulation factor X activating enzyme from Russell's viper venom, then the enzyme can be obtained, but the purity and activity are low
Solution Approach 1:
The patent applies parameter changes by optimizing chromatography conditions including pH gradients, salt concentrations, and flow rates during the purification process. The multi-step chromatography protocol with varying buffer compositions enables high-purity isolation of the enzyme while maintaining high yield, directly resolving the contradiction between manufacturing precision and productivity
Solution Approach 2:
The patent segments the purification process into multiple sequential steps: initial size exclusion chromatography, followed by anion exchange chromatography, then cation exchange chromatography, and finally affinity chromatography. This segmentation allows each step to target specific impurities, achieving both high purity and high yield that cannot be obtained with single-step methods
2Manufacturing precision
If glycosylation modification is performed during synthesis of coagulation factor X activating enzyme in vivo, then the enzyme can be produced, but the activity is affected and purity is reduced
Solution Approach 1:
The patent extracts and removes glycosylation modifications through specific enzymatic treatment and chromatography steps. By taking out the problematic glycosylation components, the patent achieves both high purity (removing modified forms) and high reliability (maintaining active enzyme), resolving the contradiction between these two parameters
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 method achieves high-purity and high-activity coagulation factor X activating enzyme suitable for large-scale industrial production, effective in stopping bleeding and treating bleeding disorders.
Implementation Method 1
subjecting the sample to anion exchange chromatography
Implementation Method 2
subjecting the sample to cation exchange chromatography
Data Source
AI summary
Provided are a novel coagulation factor X activating enzyme, a pharmaceutical composition comprising same, and use thereof in preparing a medicament for treating hemorrhage or hemorrhagic diseases. Also provided is a method for purifying a coagulation factor X activating enzyme, which adopts a sequential combination of anion exchange chromatography and cation exchange chromatography, has higher yield while ensuring the high purity and high activity of the product, greatly improves the cost-efficiency, and is beneficial to large-scale industrial production.