Peptides- Charge and Isoelectric Points, Buffering Capacity

More chemistry revision! Unfortunately this topic can be a bit of a drag...

Understand the relationship between pH, pKa and charge on ionisable amino acid side chain chains and the concept of isoelectric point. Be able to estimate net average charges on amino acid side chains at specific pHs, given the pKas of the ionisable groups. 

This was already covered in my previous post about the acid base chemistry of amino acids.

Be able to to estimate the pH range in which the isoelectric point of a peptide occurs. 

To calculate the isoelectric point of a peptide, calculate the charge on the entire peptide at different pHs by adding together the charges on each amino acid (see my previous post for how to calculate the charges of amino acids). Not sure which pHs to test? Test the pKas of the various amino acids within the peptides. The isoelectric point is the point at which the charge on the entire peptide is 0. However, you might not be able to find this point via this method- you might instead find one point where the charge is +0.5 and another point where the charge is -0.5 (or +1/-1 etc.). The pH range of the isoelectric point will lie in between these two points, but for now we haven't learned how to calculate the exact isoelectric point (though there are programs that can do this).

Understand the idea of a buffer and that buffering occurs maximally around the pKa of the ionisable group(s) of the buffer. 

Understand the concept of how added –OH reacts directly with the protonated form of a buffer in the buffering range, thus reducing the the concentration of –OH that is left to directly react with protons in solution (H+, H+(aq), H3O+) to increase the pH. This type of argument also applies to the reverse titration, starting at high pH and adding acid, where added HA directly protonates the deprotonated form of the ionisable group of the buffer rather than directly generating H+ (H+(aq), H3O+). 

A buffer is, simply put, a substance that can absorb changes in pH. Weak acids and weak bases tend to make good buffers due to their reversible dissociation equations. Here's an example for a generic acid HA:

HA + H2O <--> H3O+ + A-

Here, any acidic H+ that is added to solution simply protonates A- and has little effect on the overall solution's pH. When more basic OH- is added to solution, however, it is protonated by HA and, once again, has little effect on the solution's pH. Overall, the effect is to absorb the effects of the H+ or OH- added into the solution.

The reason why buffering occurs maximally around the pKa of the buffer is that when pH = pKa, the amount of HA is equal to the amount of A-. Hence it is equally easy for the buffer to absorb an increase in H+ and OH-.

Understand why the buffering capacity of a buffer depends on its concentration. 

This is a relatively simple concept to explain. Essentially, the more buffer that you have, the more you have to "mop up" any extra acid or base that is added to the solution.

Understand the importance of charges on proteins. 

Charge is important as it provides another way in which proteins can interact with each other. For example, proteins that interact with DNA, such as histone proteins, often have positive charges in order to interact with negatively-charged phosphate groups on the DNA.

Understand that proteins can buffer pH changes, primarily owing to the side chains of histidine residues in the proteins.

As I've mentioned before, proteins can exist in protonated and deprotonated forms, just like acids and bases. Hence, proteins can also act as buffers. I think the reason why histidine is considered important here is because the imidazole ring of histidine has several ionisable groups.