Macrocycle Polymer Removal for Hydrophobic Protein Purification
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Solution Overview
Problem
Free polymers in solutions containing hydrophobic proteins interfere with downstream processes such as affinity chromatography, mass spectrometry, and enzyme kinetic analysis, leading to significant losses and interference.
Innovation Solution
A method involving the use of macrocycles and/or hydrophobic particles to bind and remove free polymers from the solution by forming complexes, followed by separating the complexes from the solution.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If polymers are used in excess to solubilize maximum amounts of membrane protein, then the solubilization efficiency is improved, but free polymer remains in the supernatant causing interference in downstream processes
Solution Approach 1:
The patent extracts and removes free polymer from the solubilization supernatant using macrocycles (such as cyclodextrins, calixarenes, cucurbiturils, or pillararenes) that specifically bind to and sequester the free polymer, allowing it to be separated from the protein-containing solution. This resolves the contradiction by removing the harmful free polymer while preserving the solubilized proteins.
Solution Approach 2:
The patent introduces macrocycles as intermediary substances that mediate between the free polymer and the protein solution. These macrocycles act as selective binders that capture free polymer through host-guest interactions, preventing the polymer from interfering with downstream processes while allowing the proteins to remain in solution for further processing.
2Ease of manufacture
If free polymer is present in the purified solubilization supernatant, then the solubilization process is simplified, but protein binding efficiency during affinity chromatography is reduced
Solution Approach 1:
The patent applies preliminary action by removing free polymer from the solubilization supernatant before proceeding to affinity chromatography. This pre-cleaning step ensures that subsequent protein purification steps are not interfered with by free polymer, thereby maintaining high protein binding efficiency while still using a simplified solubilization approach.
3Device complexity
If free polymer remains in the solution, then additional purification steps are avoided, but downstream processes like mass spectrometry and enzyme kinetic analysis are massively interfered with
Solution Approach 1:
The patent extracts free polymer using macrocycles in a single selective binding step that can be performed in solution, avoiding the need for multiple complex purification steps like dialysis or ultrafiltration. This maintains process simplicity while effectively removing the harmful free polymer that would interfere with downstream analytical processes.
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 effectively reduces free polymer concentration by up to 95%, minimizing interference in downstream processes and preserving the integrity of hydrophobic proteins.
Implementation Method 1
contacting the solution containing the free polymers not bound to the hydrophobic protein, with a macrocycle and/or a hydrophobic particle so that the free polymers bind to the macrocycle and/or hydrophobic particle
Data Source
AI summary
A method for removing free polymers present in a solution containing a hydrophobic protein, wherein the method comprises (a) contacting the solution containing the free polymers not bound to the hydrophobic protein, with a macrocycle and/or a hydrophobic particle so that the free polymers bind to the macrocycle and/or hydrophobic particle to obtain a complex of said polymers and the macrocycle and/or hydrophobic particle, and (b) removing the complex of polymer and the macrocycle and/or hydrophobic protein from the solution. Macrocycles and a kit containing a macrocycle and/or hydrophobic particle to be employed in the method.