Question

A peptide has the following sequence: A-K-R-H-G-E-E-R (Ala-Lys-Arg-His-Gly-Glu-Glu-Arg). What is the pI for this peptide? You can use even integer approximations for pKa values:

I know the answer is 9.5 but I need to know how to get to this answer.

Answer 1

The approximate pK_a values are given. The N-terminus comes from Ala while the C-terminus is Arg.

The ionizable side chains and the pK_a values are listed below.

Amino Acid	pKa of ionizable group
Ala (N-terminus)	9.87
Lys	10.54
Arg	12.48
His	6.04
Gly	--
Glu	4.07
Glu	4.07
Arg (side chain)	12.48
Arg (C-terminus)	1.82

The pK_a values are retrieved from

https://www.elcamino.edu/faculty/pdoucette/21B-amino-acid-pka-values.pdf

Now, it is important to determine the approximate pH when the charge on the amino acid is zero. When the pH < pK_a of the ionizable group, the group is protonated (-COOH group will have charge zero and –NH₂ group will have charge +1). When pH > pK_a, the group is deprotonated (-COOH group will have charge -1 and –NH₂ group will have charge 0).

pH	Charge on								Net charge
	N-term.	Lys	Arg	His	Glu	Glu	Arg side chain	C-term.
0	+1	+1	+1	+1	0	0	+1	0	+5
1	+1	+1	+1	+1	0	0	+1	0	+5
2	+1	+1	+1	+1	0	0	+1	-1	+4
3	+1	+1	+1	+1	0	0	+1	-1	+4
4	+1	+1	+1	+1	0	0	+1	-1	+4
5	+1	+1	+1	+1	-1	-1	+1	-1	+2
6	+1	+1	+1	+1	-1	-1	+1	-1	+2
7	+1	+1	+1	0	-1	-1	+1	-1	+1
8	+1	+1	+1	0	-1	-1	+1	-1	+1
9	+1	+1	+1	0	-1	-1	+1	-1	+1
10	0	+1	+1	0	-1	-1	+1	-1	0
11	0	0	+1	0	-1	-1	+1	-1	-1
12	0	0	+1	0	-1	-1	+1	-1	-1
13	0	0	0	0	-1	-1	0	-1	-3
14	0	0	0	0	-1	-1	0	-1	-3

It is seen that the approximate pH when the net charge on the peptide is zero is 10. The approximate pI is obtained by averaging the pK_a values on either side of pH = 10. Clearly, the two pK_a values are 9.87 (N-terminus) and 10.54 (Lys side chain).

Approximate pI = ½*(9.87 + 10.54)

= 10.205

≈ 10.20 (ans).