Isoelectric Point

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nothing123

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Hi,

So glycine has a pI of approximately 5.5 (NH3+ group has a pKa of ~9 and COOH of ~2). If another amino acid that has an OH somewhere in its side chain with a pKa of ~9, would its pI be lower or higher than that of glycine? Seems pretty easy but here is where I get confused; the OH does not have a charge on it and would first act as an acid so I thought it's pI would be lower. However, apparently the pI is greater since to calculate it, we average all the pKas. However, that would be the case if the side chain was OH2+, right? Can anyone follow where I'm coming from?

Thanks.

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Hi,

So glycine has a pI of approximately 5.5 (NH3+ group has a pKa of ~9 and COOH of ~2). If another amino acid that has an OH somewhere in its side chain with a pKa of ~9, would its pI be lower or higher than that of glycine? Seems pretty easy but here is where I get confused; the OH does not have a charge on it and would first act as an acid so I thought it's pI would be lower. However, apparently the pI is greater since to calculate it, we average all the pKas. However, that would be the case if the side chain was OH2+, right? Can anyone follow where I'm coming from?

Thanks.
isoelectric point: average the pKa's of the two closest functional groups

If you stick on any functional group other than a carboxylic acid (or derivative), you're going to raise the pI. Any group you add that is not a carboxylic acid/derivative will have a higher individual pKa, and remember that alcohols have pKa of about 15.
 
What I'm getting at though is OH is more acidic than it is basic so in order to keep it neutral in the OH form, you would more likely put it in a solution that's more acidic. It just seems counterintuitive that the pI would rise since this would increase the likelihood that the H would come off leaving the O negative.
 
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What I'm getting at though is OH is more acidic than it is basic so in order to keep it neutral in the OH form, you would more likely put it in a solution that's more acidic. It just seems counterintuitive that the pI would rise since this would increase the likelihood that the H would come off leaving the O negative.
:confused:

At acidic pH and at physiological basic pH range, an alcohol behaves more like a base. When you protonate OH to form the conjugate acid, you get a pKa for the charged molecule that is less than 0, which is clearly unstable.
 
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Calculating the pI:

For neutral amino acids (all those that are not listed below as basic or acid): Pk of Carboxyl group + pK of amine group, divided by 2

For acid amino acids: (aspartate, glutamate):Add the pKs more acid, and divide by 2

For basic amino acids (lysine, arginine, histidine): add the more basic Pks, and divide by 2

I hope it helps.


My blog: www.biochemistryquestions.wordpress.com
 
:confused:

At acidic pH and at physiological basic pH range, an alcohol behaves more like a base. When you protonate OH to form the conjugate acid, you get a pKa for the charged molecule that is less than 0, which is clearly unstable.

Well you're right about that, I just thought an alcohol was a lot like water, amphoteric in nature. I'm still a little confused though. If the side chain was acidic like in glutamic acid, than the pI would go down. I understand that; to keep the second COOH group protonated, you would need a more acidic environment. The opposite applies for basic side chains. However, I find alcohols to be a little funny in that it can behave both ways (almost like having water as your side chain). If we look at the HH equation, pH = pKa + log (O-/OH), in order to keep the ratio of O-/OH low (which we want to do to keep it neutral), wouldn't we then want the pH to be as low as possible? If on the other hand the side chain was a basic NH3+, we would want the pH to be higher so the ratio of NH2/NH3+ is larger, yielding more deprotonated product. That's sort of what I'm getting at. Sorry if it seems like I'm beating a dead horse here.
 
I just thought of something interesting. Let's assume the pKa of our alcohol was in fact 15 and the pKa of our NH3+ was 9 as usual. These would be the two closest pKa's and thus would average to a pI of 12. At this pH, virtually all COOH and NH3+ would be deprotonated. Enough OH would be deprotonated as well leaving our species with a negative charge of at least -1. Do you see where I'm coming from? This is really getting confusing...
 
I just thought of something interesting. Let's assume the pKa of our alcohol was in fact 15 and the pKa of our NH3+ was 9 as usual. These would be the two closest pKa's and thus would average to a pI of 12. At this pH, virtually all COOH and NH3+ would be deprotonated. Enough OH would be deprotonated as well leaving our species with a negative charge of at least -1. Do you see where I'm coming from? This is really getting confusing...
It's probably easier to memorize the pI of the 20 common aa. Right now we're discussing an aa such as threonine, but its pI is actually 6, not 12 as we predicted. It appears that the OH group has only a modest effect on pI because it cannot hold onto a positive charge, so we don't worry about it so much...
 
Hmm, I think that might be a good idea. Strange how threonine's pI is 6.3 and serine's is 5.7. I think what one really has to know to answer a question like the one I originally posted is the pKb of the OH; then we could determine if was more likely to act as a base or an acid. If just given the pKa, all we can say is how strong the conjugate base is, not actually how basic OH is itself. Anyways, thanks for all your help.
 
I believe that the main problem is that some of you are trying to generalize Pk values of some functional groups as if they were absolute values independent of the other groups that sorround them.

Look at these two carboxyl groups:

CH3-COOH and CH2(Cl)-COOH

The Carboxyl in the 2nd group has more tendency to release the proton, since the Chloride, that is more electronegative, increase the attraction of the electrons to it, making easier to release the H+ , so this carboxyl is more acidic than the other.

Just extrapolate this knowledge, that I am sure you have, to the situation in amino acids. The OH in serine and the OH in threonine are influenced by different hydrocarbon chains, in the same way that their carboxyl and amino groups have other atoms influences, so their ability to release the protons (their acidity) is different.

In general, the pKs of carboxyl groups of amino acids are 2-4, the amino groups are around 8 to 10; the isoelectric point of the neutral aminoacids is between 5 and 7

In any fair question about calculating pI of amino acids, the proper thing is to provide in the question the Pks of the functional amino and carboxyl groups. If it happens, apply the rule I described in a former post.

I hope it helps!

My blog www.biochemistryquestions.wordpress.com
 
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there is no way you would need to know about functional groups effects on pI's for the MCAT. this is some guy in a biochem class IMO
 
there is no way you would need to know about functional groups effects on pI's for the MCAT. this is some guy in a biochem class IMO

Actually, it's an easy topic that is right in line with what they ask. For instance, what is expected when lysine, leucine, and aspartic acid are added to an DEAE-cellulose column at pH = 6.0?

By knowing that lysine is one of the three basic amino acids, we know that lysine will be positively charged at a pH = 6.0 (like amines in general). We also know that amines have high pKa values, so they make pI a bit higher than the average amino acid. Knowing that when pH < pI, it carries a positive charge tells us the same thing as intuition, told us before. So the information in the OP's question is relevant to this question. It won't bind the +-column.

By knowing that aspartic acid is one of the acidic amino acids, we know that it will be negatively charged at a pH = 6.0 (like carboxylates in general). We also know that carboxylic acids have low pKa values, so they make pI a bit lower than the average amino acid. Knowing that when pH > pI, it carries a negative charge tells us the same thing as intuition, told us before. So agian, the information in the OP's question is relevant to this question. It will bind the +-column.

There are some AAMC passages on these types of columns, so it's a good idea to have a simple way to answer these questions.
 
OH of the R group is OH portion of the carboxyl group (COOH). Deporotonation of COOH resulting in COO- is resonance stabilized. Maybe that's why you are confused on as to why OH is basic or acidic.
 

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