About the Protein Molecular Weight Calculator
This protein molecular weight calculator reads an amino acid sequence in one-letter code and returns the molecular weight of the full chain. Paste a plain sequence or a FASTA record — header lines, spaces, numbers and line breaks are ignored — and the calculator reports the average mass (what you compare with an SDS-PAGE band or use to convert mg to moles) and the monoisotopic mass (what you look for in a mass spectrum), plus the chain length and a full amino acid composition table.
It also estimates the molar extinction coefficient at 280 nm from the number of tryptophan, tyrosine and cystine residues, together with the absorbance of a 1 mg/mL solution. That is what you need to measure protein concentration on a NanoDrop or spectrophotometer using the Beer–Lambert law.
Masses are for the unmodified linear chain with a free N-terminus and C-terminus in the neutral state. Post-translational modifications, disulfide bonds (each removes about 2 Da), signal-peptide cleavage and bound cofactors are not included, so compare with experimental data accordingly.
With the default inputs, the average molecular weight is 2,395.74 Da. Change any value above to recalculate instantly.
How to use the protein molecular weight calculator
- 1Paste the protein or peptide sequence in one-letter code (FASTA headers are fine).
- 2Check the length and ignored-character count to confirm it read correctly.
- 3Use the average molecular weight for gels, molarity and mg-to-mol conversions.
- 4Use the monoisotopic mass when matching small peptides on a mass spectrometer.
- 5Use the extinction coefficient or Abs 0.1% to quantify protein at 280 nm.
Formula and method
A protein is a chain of amino acids joined by peptide bonds, each formed by losing one water molecule. Summing the residue masses (amino acid minus water) and adding a single water for the free N- and C-termini gives the mass of the chain. The average mass uses natural isotope abundances; the monoisotopic mass uses only the most abundant isotope of each element (¹²C, ¹H, ¹⁴N, ¹⁶O, ³²S), as used by ExPASy.
The extinction coefficient follows the Pace et al. (1995) method used by ProtParam: tryptophan contributes 5500, tyrosine 1490 and each disulfide-bonded cystine pair 125 M⁻¹cm⁻¹ at 280 nm in water. Abs 0.1% = ε ÷ MW is the absorbance of a 1 g/L solution.
- MW
- Average molecular weight (Da = g/mol)
- ε₂₈₀
- Molar extinction coefficient at 280 nm (M⁻¹cm⁻¹)
- n(X)
- Number of residues of amino acid X
- Abs 0.1%
- Absorbance of 1 mg/mL in a 1 cm path
Worked examples
One of each standard amino acid
The 20 residue masses sum to 2377.72 Da; adding 18.015 Da for water gives 2395.74 Da. One Trp (5500) plus one Tyr (1490) gives ε = 6990 M⁻¹cm⁻¹; the single Cys cannot form a cystine on its own.
Albumin signal peptide
The FASTA header is skipped, leaving 18 residues with an average mass of 2106.55 Da. It contains one Trp and one Tyr, so ε₂₈₀ = 6990 M⁻¹cm⁻¹.
Peptide with a possible disulfide
This oxytocin-like nonapeptide weighs 1010.19 Da as a free acid. With its two cysteines reduced ε = 1490 (one Tyr); joined as a cystine they add 125, giving 1615 M⁻¹cm⁻¹. The real hormone is cyclic and C-terminally amidated, which lowers the mass by about 3 Da.
Frequently asked questions
How do you calculate the molecular weight of a protein?+
Add up the residue mass of every amino acid in the sequence, then add 18.015 Da for one water molecule. Residue masses already exclude the water lost when each peptide bond forms.
What is the difference between average and monoisotopic mass?+
Average mass uses natural isotope abundances and matches what you weigh out or see on a gel. Monoisotopic mass uses only the lightest stable isotopes and matches the first peak of a high-resolution mass spectrum of a small peptide.
Why does my protein run at a different size on SDS-PAGE?+
Gel mobility depends on SDS binding and shape, not just mass. Glycosylation, high proline content, very acidic or basic proteins and incomplete denaturation can all shift the apparent size by 10% or more.
What is the 110 Da rule?+
It is a rough shortcut: the average amino acid residue weighs about 110 Da, so a protein of 300 residues is roughly 33 kDa. The calculator shows the exact value and the average residue mass for your sequence.
Which extinction coefficient should I use?+
Use the cystine value for proteins with disulfide bonds (most secreted and extracellular proteins) and the reduced value for intracellular proteins or samples in DTT or TCEP. The difference is small unless there are many cysteines.