Berkeley Review Isolectric Point

[Maxine450]

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I'm on the chapter with Nitrogen compounds and the isolectric point. I really don't understand the rationale of picking the pKas for multiple amino-acid compounds. In 7.26 it says that since it's written "histidine-leucine" histidine is the C-terminal and leucine is the N terminal. I understand that we want pKa 2 and 3, but WHICH ones are pKa's two and three?

For the next example on page 188, it says that the Serine is the N-terminal and isoleucine is the C-terminal, despite it being written "serine-cysteine-isoleucine". What the hell? How am I supposed to know which is the C-terminal or N-terminal?

 

[liveoak]

this is why i hate TBR. AAMC is so much simpler (but disguised as difficult)

TBR is just flat out difficult




for acidic amino acids it's pka 1 and pka 2
(coo-, nhh3+, cooh occurs at 1st equivalence point)

I always thought for basic amino acids, pi = pka2 + pka3 / 2
(coo-, nh2, nh3+ occurs at 2nd equivalent point)

 

[Maxine450]

Bump. Still need help on this..Where o where are you Berk Review Teach?

 

[raffster2]

To answer the isoelectric point questions, the order of the amino acids do not matter. You want to know the number of charged groups. If the molecule isn't cyclic, then at a low pH you will have a protonated N-terminus, and you will have protonated R groups for His, Lys, and Arg. If you have a polypeptide with a His and a Lys, for example, at low pH you would have +3 charge because of a charged NH3 terminus, charged His and charged Lys, and you would want the average of pka's 3 and 4. The order of pka's is determined by the pka's themselves. For example, pka1 would be the carboxyl terminal because it's the lowest (2-3). Next would be His, because it's around 6, then Lys and N-terminus because those are both around 9. So the isoelectric point will be when the average of pka's 3 and 4. At pka 3, C-terminus, His, and Lys are deprotonated, and at 4, N-term is also deprotonated, which will give you an average charge of 0.

The shortcut is that the charge of the polypeptide is = to the 1st pka number you're going to average. EG charge of +6 means you will average pka's 6 and 7.

 

[BilalL]

I'm on the chapter with Nitrogen compounds and the isolectric point. I really don't understand the rationale of picking the pKas for multiple amino-acid compounds. In 7.26 it says that since it's written "histidine-leucine" histidine is the C-terminal and leucine is the N terminal. I understand that we want pKa 2 and 3, but WHICH ones are pKa's two and three?

For the next example on page 188, it says that the Serine is the N-terminal and isoleucine is the C-terminal, despite it being written "serine-cysteine-isoleucine". What the hell? How am I supposed to know which is the C-terminal or N-terminal?

For 7.26 you got confused on the wording. The histidine C terminal Connects it to the N terminal of leucine. This is the peptide bond. The N terminus lies with the histidine, and the C terminus lies with the leucine on the completed peptide. So you use the pkas for the terminals on the ends if needed for the N and C terminals. Always read from left to right with the N terminus on the left and the C terminus on the right side.

So left to right serene-cysteine-isoleucine

Nterm-serine-C-N-cysteine-C-N-isoleucine-Cterm

Forgot about the inside C's and N's, they can't undergo protonation/deprotonations since they are already involved in amide bonds. However the side chains of Acidic AA's, Basic AA's, and 2 Hydrophilic AA's can (tyrosine and cysteine) and the N terminus and C terminus( the first N and last C) can.

So using the TBR trick we have to worry about 3 pka's. The cysteine, the N terminus, and the C terminus.

1. N terminus is on serine, which has a pka of 9.46.
2. C terminus on isoleucine has a pka of 2.32
3. cysteine has a pka of 8.36

The starting total charge when fully protonated is +1 since only the Nterminus is charged.

If we start at pH of 1, than the first deprotonation is on the C terminus- pka1
The next deprotonation will cysteine-pka2. The last is Nterminus-pka3

The trick TBR suggests is to take that initial total charge (+1 in this case), and use the pka of the same number and average with the next pka to get the iso electric point.

So (2.32 + 8.36)/2 = 5.34

And thats the answer.

The IE trick is actually super useful.

 

[Majik]

I'm on the chapter with Nitrogen compounds and the isolectric point. I really don't understand the rationale of picking the pKas for multiple amino-acid compounds. In 7.26 it says that since it's written "histidine-leucine" histidine is the C-terminal and leucine is the N terminal. I understand that we want pKa 2 and 3, but WHICH ones are pKa's two and three?

For the next example on page 188, it says that the Serine is the N-terminal and isoleucine is the C-terminal, despite it being written "serine-cysteine-isoleucine". What the hell? How am I supposed to know which is the C-terminal or N-terminal?

-COOH terminal has a pKa ~ 2
- R group of Histidine has a pKa ~ 8
- NH4 terminal has a pKa ~11

The lowest pKa is 1 (-COOH terminal); this will be the hydrogen that is deprotonated upon raising the pH of the solution containing the aminoacid. The next lowest is pKa 2 (Histidine in the case). This R group will contain the next Hydrogen to be deprotonated, and so on.

Another thing to keep in mind is that the -COOH group will ALWAYS have the lowest pKa. The highest pKa may vary depending on whether there are some basic amino acids in the polypeptide (I believe they have higher pKa's than the NH4+ terminal).

 

[Maxine450]

Ok I think I finally get it. I didn't realize the peptide bonds couldn't be protonated/deprotonated. It makes sense now, thanks a lot!