1. What is Protein Extinction Coefficient?

The protein extinction coefficient (ε) at 280 nm is a measure of how strongly a protein absorbs light at 280 nm wavelength. This value is crucial for determining protein concentration using UV spectrophotometry and is calculated based on the content of aromatic amino acids (tryptophan, tyrosine) and cystine residues in the protein.

2. How Does the Calculator Work?

The calculator uses the standard protein extinction coefficient equation:

Where:

• \( n_{Trp} \) — Number of tryptophan residues (molar extinction coefficient: 5500 M⁻¹ cm⁻¹)
• \( n_{Tyr} \) — Number of tyrosine residues (molar extinction coefficient: 1490 M⁻¹ cm⁻¹)
• \( n_{Cys} \) — Number of cystine (disulfide bond) residues (molar extinction coefficient: 125 M⁻¹ cm⁻¹)

Explanation: The equation sums the contributions of each chromophoric amino acid type based on their characteristic molar extinction coefficients at 280 nm.

3. Importance of Extinction Coefficient

Details: The extinction coefficient is essential for accurate protein quantification, quality control in protein purification, determination of protein concentration in various biochemical assays, and monitoring protein stability and folding.

4. Using the Calculator

Tips: Enter the number of tryptophan, tyrosine, and cystine residues from your protein sequence. All values must be non-negative integers. The calculator will compute the theoretical extinction coefficient at 280 nm.

5. Frequently Asked Questions (FAQ)

Q1: Why are only tryptophan, tyrosine, and cystine considered?
A: These three amino acids are the primary contributors to UV absorption at 280 nm due to their aromatic rings and disulfide bonds, which absorb strongly in this wavelength region.

Q2: What is the typical range for protein extinction coefficients?
A: Extinction coefficients typically range from 10,000 to 100,000 M⁻¹ cm⁻¹, depending on the protein's size and aromatic amino acid content.

Q3: How accurate is this theoretical calculation?
A: Theoretical calculations are generally accurate for most proteins, but the actual extinction coefficient can be affected by protein folding, solvent environment, and post-translational modifications.

Q4: Can I use this for protein concentration determination?
A: Yes, once you have the extinction coefficient, you can use Beer-Lambert law: Concentration = Absorbance at 280 nm / (ε × path length).

Q5: What if my protein has no aromatic amino acids?
A: Proteins without tryptophan or tyrosine will have very low extinction coefficients at 280 nm. Alternative methods like Bradford or BCA assays may be needed for concentration determination.