Is there a pH at which all proteins are negatively charged? How can it be determined?

I know that depending on the amino-acid-composition of the peptide the distance to the isoelectric point determines the charge. Is there a universal point at which all proteins are definitely negatively charged?




That depends highly on what you mean by "all." Since acid-base chemistry is equilibrium based, there is in principle always some of the less preferred species around even though it might be quite small. At the very least, there is a nonzero probability of finding the disfavored species, in your case the neutral species.


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Proteins will consist primarily of the 20 proteinogenic amino acids. Among those, lysine and arginine are positively charged at neutral $\mathrm{pH}$ while aspartate and glutamate are negatively charged. All proteins also have an amino terminus which is positively charged at neutral $\mathrm{pH}$ and an acid terminus which is negatively charged.


We don’t need to care about the acidic groups; they are already negatively charged. But if we want a $\mathrm{pH}$ level where the protein is *definitely* negatively charged, we want to deprotonate all positively charged groups.


The amino groups of the N-terminus and lysine have a $\mathrm{p}K\_\mathrm{a}\approx 10$. Thus, raising the $\mathrm{pH}$ value to $\mathrm{pH}\ 12$ should get them fully deprotonated. The guanidinium group of arginine is a little less acidic; $\mathrm{p}K\_\mathrm{a} \approx 13.6$ (approximated from the value of unsubstituted guanidinium; value found on [Wikipedia](https://en.wikipedia.org/wiki/Guanidine)). We therefore need to raise the $\mathrm{pH}$ a little bit more for certain deprotonation. If we go to the absolute limits of aquaeous $\mathrm{pH}$ of larger than $14$ (i.e. more than $1~\mathrm{M}\ \ce{NaOH}$) we should get these protonated. However, the two logarithmic units one typically assumes would mean $100~\mathrm{M}\ \ce{NaOH}$ — not possible. We can get the $\mathrm{pH}$ to around $15$, and then that’s the end of it. The fully deprotonated form should be the principal species in solution there, though.


Bear in mind that these harsh conditions are also able to hydrolyse the peptide bonds. Thus you may just destroy your protein while getting there.


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