Protein pI Calculator
Calculate the isoelectric point (pI) of a protein from its amino acid sequence.
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What it does
The protein pI calculator determines the isoelectric point of a polypeptide from its sequence. Biochemists use it to predict migration in isoelectric focusing, optimize purification buffers, and understand protein solubility and stability.
How it works
The isoelectric point (pI) is the pH at which the net charge of the protein is zero. It is found by summing the fractional charges of every ionizable group, the N-terminus, the C-terminus, and the side chains of D, E, C, Y, H, K and R, as a function of pH using their pKa values, and solving for the pH where the net charge equals zero. The tool uses the IPC 2.0 pKa values and a binary search over pH to locate the zero-charge point. The charge on each group follows the Henderson-Hasselbalch relation.
Worked example
For a simple peptide containing one Lys and one Asp plus the termini, the pI lies between the pKa values of the basic and acidic groups. A Lys-rich peptide will have a high pI near 9 to 10, while an acidic peptide rich in Asp and Glu will have a low pI near 4.
When to use it
Choose the correct pH for isoelectric focusing or two-dimensional gel separation. Decide buffer conditions that keep a protein soluble and away from its pI. Compare engineered variants to see how mutations shift the pI.
FAQ
- How is the isoelectric point of a protein calculated?
- The tool sums the charge contributed by each ionizable group at a given pH using its pKa, then finds by binary search the pH where the total net charge is zero; that pH is the pI.
- Which amino acids affect protein pI the most?
- Acidic residues Asp and Glu lower the pI, while basic residues Lys, Arg and His raise it; Cys and Tyr contribute at intermediate pH.
- What pKa values does this calculator use?
- It uses the IPC 2.0 pKa set for the ionizable side chains and termini, which gives good agreement with experimental pI for most proteins.
- Why does the net charge at pH 7 matter?
- At pH 7 you can see whether the protein is overall positive or negative under physiological conditions, which affects binding, purification and solubility.