Isoelectric Point Calculator
Isoelectric Point Results
What Is the Isoelectric Point (pI)?
The isoelectric point (pI) is the pH where an amino acid or protein has:
- Equal positive and negative charges
- A net charge of zero
- Maximum stability in many cases
At this pH, the molecule exists mainly as a zwitterion. A zwitterion has both a positive and a negative charge but balances out overall.
Why pI Matters
The isoelectric point affects:
- Protein solubility
- Enzyme activity
- Electrophoresis movement
- Protein purification methods
- Drug formulation
For example:
- At pH < pI, the molecule carries a net positive charge.
- At pH > pI, the molecule carries a net negative charge.
- At pH = pI, the net charge is zero.
Understanding this behavior helps you predict how a molecule will act in different environments.
The Basic Structure of an Amino Acid
Every amino acid contains:
- An α-carboxyl group (COOH)
- An α-amino group (NH₃⁺)
- A side chain (R group)
Each of these groups may have a pKa value. The pKa tells you at which pH that group gains or loses a proton.
Your isoelectric point calculator uses these pKa values to compute the pI.
Isoelectric Point Formula
The formula depends on the type of amino acid.
1. Neutral Amino Acids
These do not have ionizable side chains.
Formula:
pI = (pKa of α-COOH + pKa of α-NH₃⁺) / 2
You simply average the two pKa values.
2. Acidic Amino Acids
These have an extra carboxyl group in the side chain.
Examples include:
- Aspartic acid
- Glutamic acid
For acidic amino acids:
- Take the two lowest pKa values
- Average them
Formula:
pI = (pKa₁ + pKa₂) / 2
(where both pKa values are acidic)
3. Basic Amino Acids
These have an extra amino group in the side chain.
Examples include:
- Lysine
- Arginine
- Histidine
For basic amino acids:
- Take the two highest pKa values
- Average them
Formula:
pI = (pKa₂ + pKa₃) / 2
(where both pKa values are basic)
How the Isoelectric Point Calculator Works
The calculator you provided follows a simple and logical structure.
It allows users to:
- Select a predefined amino acid
- Or enter custom pKa values
- Define the side chain type (neutral, acidic, or basic)
- Click “Calculate pI”
- View step-by-step results
Input Fields
The calculator accepts:
- α-COOH pKa
- α-NH₃⁺ pKa
- Side chain pKa (optional)
- Side chain type
It also validates that all pKa values are between 0 and 14.
Step-by-Step Example Calculations
Let’s walk through three examples.
Example 1: Neutral Amino Acid (Glycine)
Glycine
Typical pKa values:
- α-COOH = 2.34
- α-NH₃⁺ = 9.60
Calculation:
pI = (2.34 + 9.60) / 2
pI = 5.97
This means glycine has no net charge at pH 5.97.
Example 2: Acidic Amino Acid (Aspartic Acid)
Aspartic acid
Typical pKa values:
- α-COOH = 1.88
- Side chain COOH = 3.65
- α-NH₃⁺ = 9.60
Use the two lowest values:
pI = (1.88 + 3.65) / 2
pI = 2.77
Aspartic acid has a much lower pI because of its extra acidic group.
Example 3: Basic Amino Acid (Lysine)
Lysine
Typical pKa values:
- α-COOH = 2.18
- α-NH₃⁺ = 8.95
- Side chain NH₃⁺ = 10.53
Use the two highest values:
pI = (8.95 + 10.53) / 2
pI = 9.74
Lysine has a high pI because it has an extra basic group.
Charge Profile Explained
Your calculator clearly shows the charge behavior:
| Condition | Net Charge |
|---|---|
| pH < pI | Positive (+) |
| pH = pI | Zero (Zwitterion) |
| pH > pI | Negative (−) |
This visual charge profile helps students quickly understand how amino acids behave in solution.
Why pKa Values Matter
The calculator includes a disclaimer, and that is important.
pKa values can change depending on:
- Temperature
- Ionic strength
- Solvent
- Local protein environment
The default values used in the calculator are standard values at 25°C in water.
For proteins with many ionizable groups, the pI becomes more complex and represents an average of many interacting pKa values.
When to Use an Isoelectric Point Calculator
You should use a pI calculator when:
- Studying amino acid chemistry
- Preparing buffer systems
- Running isoelectric focusing
- Designing protein purification experiments
- Predicting solubility behavior
It saves time and reduces calculation errors.
Benefits of This Calculator
The calculator you shared has several strengths:
- Preloaded amino acid data
- Custom input flexibility
- Input validation
- Step-by-step explanation
- Clear charge visualization
- Reset function
This makes it ideal for students, teachers, and researchers.