I'm reading about the schemes used to purify a protein from different sources. In one of these schemes, they used buffers with a pH (6.5) above the pI (5.7) of the protein to equilibrate a weak cation exchanger and elute the protein from it. The protein eluted halfway through the linear gradient. If the protein has a negative charge at that pH, why does it bind to the weak cation-exchange column?
The fact that the pH is above the pI means that the protein has a net negative charge. There may still be (and probably are) positive charges (Lys and Arg side chains and the N-terminus) on the protein surface that can interact with the cation exchanger.
Another point: pI values are often predicted, not measured, and may not be correct. Moreover, even if they are measured, they do not necessarily accurately reflect the charge on the surface of the protein, which is what interacts with the resin. Buried residues contribute to the pI, but not to the surface charge.
Cation exchanger contains negative charges which bond proteins with positive net charge . At pH higher than pI the protein has negative charge so the protein is not bond to Cation exchanger . But in one of my experiments (nearly the same conditions as yours) the protein bonds to the Cation exchanger and we found that this protein has a reagen which contain high concentration of Lys make it able to band to Cation exchanger in pH higher than its pI.
The pI is a measure of the overall charge of the protein. It does not say anything bout the charges in domains or small areas on the protein surface. The protocol you are mentioning is probably using one such pocket with positively charged residues.
A simple rule, to be certain that a protein binds to a ion exchange resin, is to use a buffer pH that is at least +/- 2 pH-units below or above the pI. When you are that far from the pI it is quite certain that the protein will bind, irrespective of local areas.
I have encountered exceptions, but they are very rare. Your example is in the grey zone.
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