would the denaturation of proteins during phenol extraction affect the binding capabilities of the proteins?
Generally protein-ligand binding will not happen if the structure of the protein changes.
But if you are trying to separate proteins which have been tagged with histidines (His tag) or thiol groups then it will work.
Hi there,
If the affinity consists in a specific native structure recognition or in using immobilized antibodies for specific interaction, the technique is useless (for instance if using GST or MBP as affinity tags or using anti Tag grafted matrix). To my knowledge only Histag is suitable for purification using Ni2+ chelating matrix in denaturing condition.
I guess we are dealing with reversible denaturation, right? Only IMAC will work with all its limitations
Depends.. see Dr Liger & Kishore's answers. If you are using a bound oligonucleotide affinity probe for a DNA binding protein then that might also work. Otherwise.. unless you can refold the protein.. forget it.
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