The pI (isoelectric point) of a peptide is found by averaging the pKa values of the two ionizable groups that flank the zwitterionic form of the peptide at its neutral point.
Here's a breakdown:
-
Identify Ionizable Groups: Determine all the ionizable groups in the peptide. These include:
- The alpha-amino group (N-terminus)
- The alpha-carboxyl group (C-terminus)
- The side chains of amino acids with acidic or basic side chains (e.g., Asp, Glu, His, Lys, Arg, Tyr, Cys)
-
Assign pKa Values: Look up or estimate the pKa values for each ionizable group. Typical pKa values can be found in biochemistry textbooks or online resources. Remember that the specific chemical environment within the peptide can slightly alter the pKa values compared to those of free amino acids.
-
Determine the Relevant pKa Values: To find the pI, you need the two pKa values that define the pH range where the peptide is neutral (zwitterionic). This involves considering the protonation states of the ionizable groups.
-
If the peptide has no acidic or basic side chains, average the pKa of the N-terminus and the C-terminus.
-
If the peptide contains acidic side chains (Asp, Glu), average the two lowest pKa values, corresponding to the two acidic groups that must be protonated for the peptide to be neutral.
-
If the peptide contains basic side chains (Lys, Arg, His), average the two highest pKa values, corresponding to the two basic groups that must be deprotonated for the peptide to be neutral.
-
-
Calculate the Average: Add the two relevant pKa values together and divide by two. This average is the pI.
Example:
Let's say a peptide has the following ionizable groups and estimated pKa values:
- N-terminus: pKa = 9.0
- C-terminus: pKa = 2.0
- Lysine side chain: pKa = 10.5
To find the pI, we average the two highest pKa values (the basic groups), which are 9.0 and 10.5.
pI = (9.0 + 10.5) / 2 = 9.75
Therefore, the pI of this peptide is approximately 9.75.
