Acid base Chenistry Homework help!?
Histidine was electrophoresed in polyacrylamide at pH 6.0 with alanine (pI=6.0) and arginine (pka's 2.2, 9.0, and 12.5). At the end of the experiment, arginine had migrated toward the cathode 6.0 cm, alanine was still at the origin, and histidine was present at 3.0 toward cathode. No histine was present at 0.0 or 6.0 cm. Explain how is it possible that the histidine molecules behaved as though they had a fraction of one electronic charge, when they actually have either +1 or 0 charge.
Jane Protein thinks this result is bogus (histidine should be present only at 0.0 and 6.0 cm) and so this result shows a major experimental error but John Biochem says this result is exactly what any student who understands acid-base equilibrium would predict. Decide whether you agree with Jane or John and explain why.
My comment:
Well, I think that the side chain, on histidine can both donate and accept protons. The imidazole side chain has two nitrogens with different properties: One is bound to hydrogen and donates its lone pair to the aromatic ring and as such is slighty acidic, whereas the other one donates only one electron pair to the ring so it has a free lone pair and is basic. Is this true for PH=6?
Yes, you are correct. The side chain of histidine contains an imidazole group, which has two nitrogen atoms with different properties. At pH 6.0, the side chain of histidine can be in a state where one nitrogen is positively charged and the other is neutral. This is because at pH 6.0, the imidazole group is partially protonated, meaning that one of the nitrogens has gained a hydrogen ion (H+) and carries a positive charge, while the other nitrogen remains neutral. This results in histidine having a net positive charge of +1.
Now, let's analyze the given information about the electrophoresis experiment. Alanine has a pI (isoelectric point) of 6.0, which means it will have no net charge at pH 6.0 and will not migrate in the electric field. Therefore, it is expected that alanine remains at the origin.
Arginine, on the other hand, has multiple pKa values (2.2, 9.0, and 12.5), indicating multiple ionizable groups. At pH 6.0, arginine will be predominantly in a state where all its ionizable groups are partially protonated. This means that at pH 6.0, arginine will have a net positive charge of +1. Since it migrates toward the cathode 6.0 cm, it suggests that it is attracted to the negatively charged electrode due to its positive charge.
Now, let's consider histidine. At pH 6.0, histidine will have a net charge of +1 due to the partially protonated imidazole side chain. However, based on the given information, histidine was found at 3.0 cm toward the cathode, meaning it migrated only halfway to the position of arginine.
This can be explained by the fact that histidine can act as a zwitterion, which means it can exist in a neutral form with no net charge. The neutral form of histidine may not move as far as the positively charged arginine due to differences in size and shape, as well as interactions with the gel matrix used in the electrophoresis.
To address the disagreement between Jane and John, it is important to note that the observed migration of histidine does not indicate a major experimental error. Instead, it reflects the behavior expected from a histidine molecule with both positive and neutral charges. John Biochem's understanding of acid-base equilibrium predicts this behavior, whereas Jane Protein's skepticism may stem from a lack of knowledge about histidine's properties and the concept of zwitterions.
Overall, I agree with John Biochem that the result observed in the experiment is consistent with the acid-base equilibrium properties of histidine and the behavior of amino acids during electrophoresis.