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🧬

Isoelectric Point: Protein pI and Amino Acids

The isoelectric point (pI) is the pH at which a molecule has zero net charge. For amino acids: simple pI = (pKa₁ + pKa₂)/2; acidic side chain pI = (pKa₁ + pKa₃)/2; basic side chain pI = (pKa₂ + pKa₃)/2. Essential for protein purification, electrophoresis, and solubility prediction.

Concept Fundamentals
—
pI
—
Charge pH 7
—
Side Chain
pH = pI
Zwitterion
Calculate Isoelectric PointAmino acid pI | Zwitterion | Electrophoresis

Why This Chemistry Calculation Matters

Why: pI determines protein solubility, electrophoresis mobility, and purification strategies. At pI, proteins precipitate; away from pI they are charged and soluble.

How: Select amino acid from database or enter custom pKa values. Simple AA: pI = (pKa₁+pKa₂)/2. Acidic (Asp, Glu): pI = (pKa₁+pKa₃)/2. Basic (Lys, Arg, His): pI = (pKa₂+pKa₃)/2.

  • ●At pI, net charge = 0; minimum solubility.
  • ●Isoelectric focusing separates proteins by pI.
  • ●pI affects drug solubility and membrane permeability.
Sources:IUPAC Gold BookExPASy ProtParam
🧬Isoelectric Point (pI)pI = (pKa₁ + pKa₂) / 2 | Zwitterion

Compact Examples

🧬 Glycine (Simple)
Simplest amino acid - pKa1=2.34, pKa2=9.60, pI=5.97
⚡ Glutamic Acid (Acidic)
Acidic side chain - pKa3=4.25, pI=3.22
➕ Lysine (Basic)
Basic side chain - pKa3=10.53, pI=9.74
🔬 Histidine (Special)
Imidazole side chain - pKa3=6.00, pI=7.59
🌿 Tyrosine (Aromatic)
Phenolic side chain - pKa3=10.07, pI=5.66
🔗 Cysteine (Thiol)
Sulfhydryl group - pKa3=8.18, pI=5.07
⚡ Aspartic Acid
Acidic side chain - pKa3=3.65, pI=2.77
➕ Arginine
Guanidinium group - pKa3=12.48, pI=10.76
📝 Custom Simple AA
Calculate pI from pKa1 and pKa2
📝 Custom Charged AA
Calculate pI with side chain pKa

Inputs

For reference (pKa values are temperature-dependent)
Enter full name, 3-letter code, or 1-letter code

For educational and informational purposes only. Verify with a qualified professional.

🔬 Chemistry Facts

🧬

Glycine pI = 5.97; Glutamate pI = 3.22; Lysine pI = 9.74.

— Biochemistry

⚡

At pI, proteins have minimum solubility—used for precipitation.

— Protein chem

🔬

Isoelectric focusing separates proteins by pI in a pH gradient.

— Electrophoresis

📐

Zwitterion: NH₃⁺-R-COO⁻ at pI; charge = 0.

— IUPAC

📋 Key Takeaways

  • • pI = pH at which net charge = 0 (zwitterion)
  • • Simple AA: pI = (pKa₁ + pKa₂) / 2
  • • Acidic side chain: pI = (pKa₁ + pKa₃) / 2
  • • Basic side chain: pI = (pKa₂ + pKa₃) / 2

Did You Know?

🧬

Glycine pI = 5.97; Glutamate pI = 3.22; Lysine pI = 9.74.

Source: Biochemistry

⚡

At pI, proteins have minimum solubility—used for precipitation.

Source: Protein chem

🔬

Isoelectric focusing separates proteins by pI in a pH gradient.

Source: Electrophoresis

📐

Zwitterion: NH₃⁺-R-COO⁻ at pI; charge = 0.

Source: IUPAC

💊

pI affects drug solubility and membrane permeability.

Source: Pharma

🌡️

pI is temperature-dependent; values often at 25°C.

Source: CRC Handbook

How the Isoelectric Point Calculator Works

Select an amino acid from the database or enter custom pKa values. pI is the average of the two pKa values that bracket the neutral charge state.

Simple

pI = (pKa₁ + pKa₂) / 2

Charged

Acidic: (pKa₁+pKa₃)/2; Basic: (pKa₂+pKa₃)/2

Expert Tips

Database

Use 20 standard amino acids with built-in pKa values.

Acidic vs Basic

pKa₃ < 7: acidic; pKa₃ > 7: basic.

IEF

Isoelectric focusing separates by pI.

Solubility

Minimum solubility at pI.

Amino Acid pI Reference

AACodepIType
GlycineG5.97nonpolar
AlanineA6.01nonpolar
ValineV5.97nonpolar
LeucineL5.98nonpolar
IsoleucineI6.02nonpolar
MethionineM5.75nonpolar
ProlineP6.29special
PhenylalanineF5.48aromatic

Frequently Asked Questions

What is pI?

pH at which net charge = 0. Zwitterion form.

How to calculate pI for Glycine?

pI = (2.34 + 9.60) / 2 = 5.97.

Acidic vs basic side chain?

Acidic (Asp, Glu): pI = (pKa₁+pKa₃)/2. Basic (Lys, Arg): pI = (pKa₂+pKa₃)/2.

What is zwitterion?

NH₃⁺-R-COO⁻; both + and - charges, net 0.

Why minimum solubility at pI?

No net charge ⇒ no electrostatic repulsion ⇒ aggregation.

Isoelectric focusing?

Separates proteins by pI in pH gradient.

pI vs pKa?

pKa = single group; pI = whole molecule at zero charge.

Key Numbers

5.97
Glycine pI
3.22
Glutamate pI
9.74
Lysine pI
7.59
Histidine pI

📚 Official Sources

IUPAC Gold Book ↗
Isoelectric point definition
UniProt Database ↗
Protein pI data
ExPASy ProtParam ↗
Protein analysis tools

⚠️ Disclaimer: pI values are temperature-dependent. For proteins, pI depends on all ionizable groups. Consult IUPAC and ExPASy ProtParam for authoritative data.

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