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Lesson 7 BCH_part 8 (Histidine in a gel electrophoresis)

Posted: Fri Feb 14, 2020 12:28 pm
by sen117

I am confused with the following question in lesson 7 video:

Q. In an electrophoresis gel with a pH=9.5, match the AA to its overall charge or node to which it will migrate
A. His (- charge)

I thought His can either be charged + when pH <6 or be uncharged. At pH greater than 7.6 (PI of His), does His lose H+ from imidazole resulting in it's N bearing a negative charge? If so, the categorization of AA (such as His being a basic + charged AA), is only true at physiological pH?

Thank you for your help in advance!

Re: Lesson 7 BCH_part 8 (Histidine in a gel electrophoresis)

Posted: Wed Feb 19, 2020 6:40 pm
by NS_Tutor_Mathias
This is an exceptionally oddly constructed question. There are a few big lessons to be learned here:
1. Do not try to confuse concepts the way it is being done here, like isoelectric point, anode and cathode migration and protonation states. That will just make a mess in your head.

2. There is a difference between the idea of what species exist in solution at any point in time and what the net charge of the species in solution is. That is to say, if I have more negatively charged species than positively charged, I must be past the isoelectric point - but that would NOT mean that the majority of that particular molecule would be negatively charged. The majority may be neutral, as is the case here with histidine.

3. For basic AAs (Histidine, Lysine, Arginine), the pKa commonly given is the pKa of the conjugate acid. So the common form of histidine in a pH 9.5 solution is the neutral form - however, the fully deprotonated form (at both the N and C terminus) outnumbers the protonated form past the pI, so the net charge of all the histidine in solution is ever so slightly negative.

And finally, it would indeed migrate to the anode - however, not as far as more basic AAs might.