Protein Molecular Weight Calculator
A protein's mass is the sum of its residues plus one water. The protein molecular weight calculator does the sum from Expasy's residue masses, and the extinction coefficient alongside, so a reading at 280 nm becomes a concentration.
Molecular weight
- Average molecular weight The sum of 129 residues at average isotopic masses, plus one water (18.01524 Da) for the two chain ends. 1 Da is 1 g/mol, so this is also 14,313.14 g/mol, the figure for weighing out protein or reading an SDS-PAGE ladder.
- 14,313.14 Da (14.31 kDa)
- Monoisotopic mass Every atom as its most abundant isotope, the mass a mass spectrometer resolves as the first peak of a small peptide. For a protein of more than a few kDa that peak is too small to see and the average mass is the useful one.
- 14,303.88 Da
- Length 110.95 Da per residue on average. The rule of thumb of 110 Da per amino acid comes from exactly this division over typical proteins.
- 129 residues
- If the cysteines pair (4 disulfides) Each disulfide bond removes two hydrogens, 2.02 Da. The figure above assumes free thiols, as Expasy's tools do; whether the bonds form depends on the protein, not the sequence.
- 14,305.08 Da
- Extinction coefficient, 280 nm 6 Trp x 5,500 + 3 Tyr x 1,490 + 4 cystine x 125. With every cysteine reduced it would be 37,470. For the native protein in water.
- 37,970 M^-1 cm^-1
- Abs 0.1% (1 g/L) The absorbance of a 1 mg/mL solution over 1 cm: the extinction coefficient divided by the molecular weight. Proteins differ by a factor of several here, which is why A280 = 1 does not mean 1 mg/mL.
- 2.653
- Concentration at A280 of 1 Beer-Lambert: concentration = A / (extinction coefficient x path), with a 1 cm path. Subtract the buffer blank first; anything else in the sample that absorbs at 280 nm, such as nucleic acid, inflates it.
- 0.377 mg/mL (26.34 µM)
| Amino acid | Count | Mol % | Mass in chain (Da) |
|---|---|---|---|
| Alanine (Ala, A) | 12 | 9.3% | 852.95 |
| Arginine (Arg, R) | 11 | 8.5% | 1,718.06 |
| Asparagine (Asn, N) | 14 | 10.9% | 1,597.45 |
| Aspartic acid (Asp, D) | 7 | 5.4% | 805.62 |
| Cysteine (Cys, C) | 8 | 6.2% | 825.11 |
| Glutamine (Gln, Q) | 3 | 2.3% | 384.39 |
| Glutamic acid (Glu, E) | 2 | 1.6% | 258.23 |
| Glycine (Gly, G) | 12 | 9.3% | 684.62 |
| Histidine (His, H) | 1 | 0.8% | 137.14 |
| Isoleucine (Ile, I) | 6 | 4.7% | 678.96 |
| Leucine (Leu, L) | 8 | 6.2% | 905.28 |
| Lysine (Lys, K) | 6 | 4.7% | 769.04 |
| Methionine (Met, M) | 2 | 1.6% | 262.39 |
| Phenylalanine (Phe, F) | 3 | 2.3% | 441.53 |
| Proline (Pro, P) | 2 | 1.6% | 194.23 |
| Serine (Ser, S) | 10 | 7.8% | 870.78 |
| Threonine (Thr, T) | 7 | 5.4% | 707.74 |
| Tryptophan (Trp, W) | 6 | 4.7% | 1,117.28 |
| Tyrosine (Tyr, Y) | 3 | 2.3% | 489.53 |
| Valine (Val, V) | 6 | 4.7% | 594.80 |
- Post-translational modifications are not included: glycosylation, phosphorylation, a removed initiator methionine, a cleaved signal peptide or an acetylated N-terminus all change the real mass, and glycans can add many kDa. Enter the mature sequence for the chain you actually have.
- Residue masses and the extinction constants are Expasy's (SIB), and the sums reproduce Expasy's Compute pI/Mw and ProtParam to the two decimals they print.
How the Protein Molecular Weight Calculator works
A protein's molecular weight is the sum of its residue masses plus the mass of one water molecule: MW = sum of residues + 18.02 Da. A residue is an amino acid less the water it gave up to form peptide bonds, so a chain of n amino acids has lost n - 1 waters, and adding one back for the free ends gives the right total. The insulin A chain, GIVEQCCTSICSLYQLENYCN, is 21 residues summing to 2,365.69 Da; add 18.02 and its molecular weight is 2,383.71 Da, which is exactly what Expasy's Compute pI/Mw reports for the same sequence.
Average or monoisotopic. The average mass uses each element's natural mix of isotopes and is the right figure for anything weighed in bulk: making up a solution, reading a gel, working out moles. The monoisotopic mass assumes every atom is its lightest common isotope, and it is what a mass spectrometer resolves as the first peak of a small peptide. For a peptide of a few residues the two differ by a fraction of a dalton; for lysozyme, the default here, they differ by about 9 Da, and as proteins grow the monoisotopic peak becomes too small to observe at all. Quoting the monoisotopic mass of a 50 kDa protein is a common mix-up between the two.
What the sequence leaves out. The mass is of the chain as written. A signal peptide that is cleaved, an initiator methionine that is removed, disulfide bonds (each takes away two hydrogens, about 2.02 Da, which the page shows separately), phosphates, acetyl groups and glycans are not in it. Glycosylation in particular can add many kilodaltons, and it is one reason a protein runs at a different apparent size on SDS-PAGE than its sequence predicts. Enter the mature chain you actually have; the default is lysozyme's 129-residue mature chain, without the 18-residue signal peptide UniProt lists in front of it.
Letters that are not residues. B means Asp or Asn, Z means Glu or Gln, J means Leu or Ile, and X means any amino acid. None has a single mass, so they are counted, reported and left out rather than guessed, and the page gives the range the true mass would fall in. Spaces, digits and a FASTA header are stripped and counted. The form takes one line, so a pasted FASTA record often arrives with its header joined to the sequence; the header is taken to end at its last punctuation mark, and the page says when it had to do that.
The extinction coefficient turns a spectrophotometer reading into a concentration. At 280 nm almost all of a protein's absorbance comes from tryptophan, tyrosine and cystine, so ProtParam's method estimates it as 5,500 per Trp, plus 1,490 per Tyr, plus 125 per cystine, in M^-1 cm^-1 for the native protein in water. Lysozyme has 6 Trp, 3 Tyr and 4 disulfides: 33,000 + 4,470 + 500 = 37,970. Divide by the molecular weight and you have the absorbance of a 1 mg/mL solution, 2.653 for lysozyme, so a reading of 1.0 is 0.377 mg/mL rather than the 1 mg/mL people often assume. Free cysteines barely absorb, which is why the figure depends on whether they are paired; choose the case you have. Proteins without tryptophan can be off by more than 10%.
The calculation is the one Expasy's tools use, from the same residue masses, and it reproduces their published numbers to the two decimals they print.
Protein Molecular Weight Calculator FAQ
How do you calculate the molecular weight of a protein?
Add the residue mass of every amino acid in the sequence, then add one water (18.02 Da average) for the free amino and carboxyl ends. Residue masses are each amino acid minus one water, because forming each peptide bond releases a water. The insulin A chain comes to 2,383.71 Da.
Is 110 Da per amino acid accurate?
As a rule of thumb for typical proteins, yes: dividing a protein's mass by its length usually gives something near 110 Da. For a particular sequence it can be well off, since residues range from glycine at 57 Da to tryptophan at 186 Da. Use the sum when the number matters.
What is the difference between average and monoisotopic mass?
Average mass uses each element's natural isotope mix and is what you weigh. Monoisotopic mass uses only the most abundant isotope of each element and is the first peak a mass spectrometer resolves for a small peptide. For large proteins that peak is too small to observe and the average mass is used.
Does this include glycosylation and other modifications?
No. Post-translational modifications, disulfide bonds, removed signal peptides and initiator methionines all change the real mass and none is in the sum. The page shows what disulfides would subtract; for anything else, add the modification's mass yourself or measure it.
How do I get protein concentration from A280?
Divide the absorbance by the extinction coefficient and the path length to get molar concentration, or by the Abs 0.1% figure to get mg/mL. Enter your A280 above and the page does both, using the sequence's own coefficient rather than assuming 1.0 per mg/mL.
Sources
- SIB Swiss Institute of Bioinformatics, Expasy 'Protein Identification Tools - Mass values used', section 2 'Amino acids' (standard and non-standard) and section 3 'Other mass values' (H, H2O). Method from the Compute pI/Mw documentation: 'Protein Mw is calculated by the addition of average isotopic masses of amino acids in the protein and the average isotopic mass of one water molecule.' (2026)
- SIB Expasy, 'ProtParam - Documentation', section 'Extinction coefficients' (excerpt from Gasteiger et al., Protein Identification and Analysis Tools on the Expasy Server, The Proteomics Protocols Handbook, Humana Press 2005). The page describes the method as Edelhoch's (1967), validated for native proteins by Gill and von Hippel (1989) Anal. Biochem. 182(2):319-326, with the Trp and Tyr coefficients of Pace, Vajdos, Fee, Grimsley and Gray (1995) Protein Sci. 4(11):2411-2423. (2026)