Since acetic acid is considered a weak acid (strong within the scale of weak acids), there is a range at which the acetic acid and acetate will form a stable buffer.
Using the Henderson-Hasselbalch equation for buffers, it’s possible to calculate the pH based on the ratio of acid to conjugate base, or calculate the ratio given the pH and pKa value. Finding charge on a protonated amine:
The pKa value given for the amino group on any amino acid specifically refers to the equilibrium between the protonated positive nitrogen and deprotonated neutral nitrogen. You’ll never see a neutral nitrogen deprotonated to form a negative on an amino acid.
The pKa of the protonated methylamine conjugate acid is like this: at physiological pH, there is an …show more content…
So how does this relate to the isoelectric point? Do we randomly pick a value?
The answer is no, but let’s take a closer look at the pKa values first.
As explained in the buffer video above , when the pH is exactly at the pKa value, we have an ideal buffer where the molecules exist in equilibrium. For example, if pH = 2.34, which is the pKa of the carboxyl group, what is the net charge?
Since this is the carboxyl buffer zone, we’ll have 50% neutral molecules where the carboxyl is deprotonated, and 50% positive molecules where the carboxyl is protonated.
Now if we raise the pH to 9.60, the pKa of the protonated amino group, we get yet another buffer.This time there is an equilibrium between the protonated neutral zwitterion and the deprotonated negative molecule; once again in a 50/50 ratio.
So if each pKa value gives us a 50% neutral molecule, and the isoelectric point is the pH of exact neutrality, we need to go EXACTLY halfway between the two values that give us 50% neutral. The first value gives us 50% neutral and 50% +1. The second value gives us 50% neutral and 50% -1.
The isoelectric point is the average of the 2 pKa values that have a neutral molecule as one of its equilibrium …show more content…
Charge determination function:
The process of calculating the charge on the protein at a particular pH is handled by the charge de termination function. In essence, the charge of the protein is equivalent to the sum of the fractional charges of the protein’s charged groups: Z = Nterm + Cterm + α∗K + β ∗R + γ ∗H + δ∗D + ∗E + ζ ∗C + η∗Y, where Nterm, Cterm, K, R, H, D, E, C, and Y are the charges these groups take on at a particular pH and the Greek letters before them are the count of each amino acid residue from the protein sequence. Z, the sum of these terms, is the charge on the entire protein.
This summation places some limitations on the accuracy of this algorithm. First, by adding together the charges in this way, the algorithm assumes that each group’s charge is independent of all others; if the protein contains an arginine side chain and an aspartic acid side chain, for example, these two groups are assumed to take on a charge irrespective of their locations in the protein sequence. If a basic residue is adjacent to an acidic residue, each probably does change the other’s ability to take on a charge, but this effect is ignored in this algorithm
Individual charge dtermination
Using the Henderson-Hasselbalch equation for buffers, it’s possible to calculate the pH based on the ratio of acid to conjugate base, or calculate the ratio given the pH and pKa value. Finding charge on a protonated amine:
The pKa value given for the amino group on any amino acid specifically refers to the equilibrium between the protonated positive nitrogen and deprotonated neutral nitrogen. You’ll never see a neutral nitrogen deprotonated to form a negative on an amino acid.
The pKa of the protonated methylamine conjugate acid is like this: at physiological pH, there is an …show more content…
So how does this relate to the isoelectric point? Do we randomly pick a value?
The answer is no, but let’s take a closer look at the pKa values first.
As explained in the buffer video above , when the pH is exactly at the pKa value, we have an ideal buffer where the molecules exist in equilibrium. For example, if pH = 2.34, which is the pKa of the carboxyl group, what is the net charge?
Since this is the carboxyl buffer zone, we’ll have 50% neutral molecules where the carboxyl is deprotonated, and 50% positive molecules where the carboxyl is protonated.
Now if we raise the pH to 9.60, the pKa of the protonated amino group, we get yet another buffer.This time there is an equilibrium between the protonated neutral zwitterion and the deprotonated negative molecule; once again in a 50/50 ratio.
So if each pKa value gives us a 50% neutral molecule, and the isoelectric point is the pH of exact neutrality, we need to go EXACTLY halfway between the two values that give us 50% neutral. The first value gives us 50% neutral and 50% +1. The second value gives us 50% neutral and 50% -1.
The isoelectric point is the average of the 2 pKa values that have a neutral molecule as one of its equilibrium …show more content…
Charge determination function:
The process of calculating the charge on the protein at a particular pH is handled by the charge de termination function. In essence, the charge of the protein is equivalent to the sum of the fractional charges of the protein’s charged groups: Z = Nterm + Cterm + α∗K + β ∗R + γ ∗H + δ∗D + ∗E + ζ ∗C + η∗Y, where Nterm, Cterm, K, R, H, D, E, C, and Y are the charges these groups take on at a particular pH and the Greek letters before them are the count of each amino acid residue from the protein sequence. Z, the sum of these terms, is the charge on the entire protein.
This summation places some limitations on the accuracy of this algorithm. First, by adding together the charges in this way, the algorithm assumes that each group’s charge is independent of all others; if the protein contains an arginine side chain and an aspartic acid side chain, for example, these two groups are assumed to take on a charge irrespective of their locations in the protein sequence. If a basic residue is adjacent to an acidic residue, each probably does change the other’s ability to take on a charge, but this effect is ignored in this algorithm
Individual charge dtermination