Isoelectric Point (pI)

The isoelectric point is the pH at which a peptide's positive and negative charges exactly cancel, leaving zero net charge. It is computed from the dissociation constants of the ionisable groups present, with the terminal amine near pH 9 and the terminal carboxyl near 3, aspartate and glutamate near 4, histidine near 6, tyrosine near 10, lysine near 10.5 and arginine near 12.5, by finding the pH at which the summed charges reach zero.

It matters because electrostatic repulsion is what keeps like molecules apart in solution, and at the isoelectric point there is none, so solubility usually reaches a minimum and aggregation a maximum. Insulin glargine turns this into a mechanism: two arginines added to the B chain and an asparagine-to-glycine change on the A chain shift the value from about 5.4 towards neutral, so the drug is soluble in its acidic vial and precipitates as a microcrystalline depot on injection at physiological pH.

Formulation therefore works away from the isoelectric point, usually by a pH unit or two, and the same figure dictates analytical choices: which ion-exchange chemistry will retain the peptide, how it migrates in isoelectric focusing, and which surfaces it adheres to at a given buffer pH. Knowing it before any experiment tells you where solubility trouble will appear.

A calculated value is an estimate and is regularly treated as a measurement. Tabulated dissociation constants are those of free amino acids, whereas in a folded peptide a buried residue or a neighbouring charge can shift a side-chain constant by more than a unit. It also predicts only the electrostatic contribution to solubility: a peptide can be poorly soluble far from its isoelectric point for hydrophobic reasons, and a well-designed formulation can be stable near it.