The protein molecular weight calculator above sums the average residue masses of every amino acid in a sequence, adds the mass of one water molecule for the free ends, and reports the result in daltons and kilodaltons. Alongside it you get the residue count, the predicted molar extinction coefficient at 280 nm, the absorbance a 1 mg/mL solution would give, the number of moles in a mass you specify, and a breakdown of the sequence by residue class.
Arb Digital publishes this as a free bench tool that runs entirely in your browser, so a sequence you paste never leaves your machine. It is aimed at the everyday questions — how many moles are in this tube, what should this band run at, what coefficient do I put into a concentration calculation — rather than at proteomics-grade mass prediction.
What This Protein Molecular Weight Calculator Does
A polypeptide is formed by condensation: each peptide bond releases one water molecule, so the chain weighs less than the sum of its free amino acids. The clean way to handle that is to work in residue masses, which are the amino acid masses already less water, and then add a single water back for the two free termini. That is what this page does, using average isotopic masses rather than monoisotopic ones.
The extinction coefficient uses the same method the ExPASy ProtParam documentation describes: the number of tyrosines multiplied by 1,490, plus tryptophans multiplied by 5,500, plus cystines multiplied by 125. Selecting the oxidised cysteine state pairs the cysteines into cystines and includes their contribution; the reduced state leaves them out, because free cysteine absorbs negligibly at 280 nm.
Those three per-residue constants were calibrated experimentally in How to measure and predict the molar absorption coefficient of a protein, published by Pace and colleagues in Protein Science in 1995, which compared predicted coefficients against measured values across a large set of proteins. That calibration is the reason coefficients from independent tools agree closely, and it is also why the prediction is described as applying to a folded protein in water rather than to a denatured one.
Boundary worth stating: this page parses a peptide sequence. If you need the molar mass of an inorganic compound written as a chemical formula, that is a different parser and a different job — use our molar mass calculator for those.
How to Use It
- Paste the sequence in single-letter code. FASTA headers, line breaks, numbers and whitespace are stripped automatically.
- Set the cysteine state to match how your protein exists in the buffer you are measuring in. It changes the extinction coefficient, not the mass.
- Set the chain count if you want the mass of a homodimer or higher multimer rather than a single subunit.
- Enter a mass in milligrams to convert straight to moles, which is usually the reason you wanted the molecular weight in the first place.
- Check the unrecognised character count in the sub-line. A non-zero value means something in your paste was not a standard residue.
The Formula and How It's Calculated
Molecular weight is the sum of the average residue masses plus 18.01524 daltons for water. Taking a 20-residue test peptide containing exactly one of each standard amino acid, the residue masses total 2,377.72 daltons, and adding water gives 2,395.74 daltons, or 2.40 kilodaltons. That sequence contains one tryptophan and one tyrosine, so its extinction coefficient is 5,500 + 1,490 = 6,990 M⁻¹cm⁻¹, and the absorbance of a 1 mg/mL solution is 6,990 ÷ 2,395.74 = 2.918.
Moles follow from mass divided by molecular weight. One milligram of that peptide is 0.001 ÷ 2,395.74 = 4.174 × 10⁻⁷ moles, which the grid reports as 0.417 micromoles. If you set the chain count above one, every downstream figure — mass, coefficient, moles — scales with it, since a homomultimer is simply the same sequence repeated.
Average Mass Versus Monoisotopic Mass
Elements exist as mixtures of isotopes, and there are two defensible ways to weigh a molecule made of them. Average mass uses the natural abundance-weighted mass of each element, which is what a balance measures and what any bulk calculation needs. Monoisotopic mass uses only the most abundant isotope of each element, which is what a high-resolution mass spectrometer resolves for a small molecule.
This calculator reports average mass, because that is the correct figure for converting between grams and moles. If you are matching a peak in a high-resolution spectrum, monoisotopic mass is the right comparison and it is measurably lower — the gap widens with size, reaching more than a dalton for a modest peptide and far more for a whole protein. Comparing an average mass to a monoisotopic peak is a common and confusing source of apparent discrepancy.
Why Your Protein Never Runs Where the Sequence Says
A predicted molecular weight describes the bare translated chain, and very few proteins stay that way. Signal peptides are cleaved after targeting, N-terminal methionine is frequently removed, and propeptides are excised during maturation, all of which reduce the mass. Post-translational modification moves it the other way: N-linked glycosylation can add tens of kilodaltons, and phosphorylation adds 80 daltons per site.
Electrophoretic mobility adds its own layer. SDS-PAGE separates by the amount of detergent bound, which correlates with length but is distorted by unusually acidic or basic sequences, by proline-rich regions and by residual structure. Highly acidic proteins routinely run slower than their true mass, sometimes by ten per cent or more. A band that does not sit where you predicted is therefore not evidence that the calculation is wrong, and the composition bars on this page will often tell you why.
Tags, Linkers and the Construct You Actually Have
The most frequent error with a tool like this is calculating the mass of the wrong sequence. A hexahistidine tag, a thrombin or TEV cleavage site, a linker, a fluorescent fusion partner and the residual scar left after cleavage are all real mass that belongs in the number if they are present in your construct. Paste the full expressed sequence, including the tag, rather than the sequence from the database entry.
It matters most for the extinction coefficient. A fusion partner rich in tryptophan can dominate the absorbance of the whole construct, so using the coefficient of the untagged protein to quantify a tagged one produces a systematically wrong concentration. Once you have the right coefficient, our protein concentration calculator converts an A280 reading into mg/mL using it.
Reading the Composition Breakdown
The four bars group residues by side-chain character: nonpolar, polar but uncharged, acidic, and basic. They are a quick sanity check rather than a structural prediction. A soluble globular protein typically shows a substantial nonpolar fraction buried in its core, while membrane-spanning sequences skew heavily nonpolar across the whole chain.
The balance between acidic and basic residues is the more directly useful comparison, because it is what drives the isoelectric point and therefore behaviour on ion exchange chromatography. A sequence with a clear excess of aspartate and glutamate over lysine, arginine and histidine will carry net negative charge at neutral pH and bind an anion exchanger. This page does not compute a pI, since that needs a full charge-state model rather than a residue count, but the bars show you which way the sequence leans.
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Browse All Free Tools Contact Arb DigitalCommon Mistakes to Avoid
- Calculating the untagged sequence when the protein you have carries a tag, linker or fusion partner.
- Comparing this average mass against a monoisotopic peak from a high-resolution mass spectrometer.
- Assuming a gel band at the wrong height means an error, when mobility is distorted by charge, proline content and modification.
- Leaving the cysteine state on reduced for a disulphide-bonded protein, which understates the extinction coefficient.
- Including the mass of a signal peptide that is cleaved before the mature protein ever exists.
Related Free Tools From Arb Digital
Pair this with the protein concentration calculator, which uses the extinction coefficient and molecular weight computed here. For chemical formulas rather than sequences there is the molar mass calculator, and for solution work the molarity calculator. On the nucleic acid side, the DNA to mRNA converter handles transcription, the primer melting temperature calculator covers PCR design, and the DNA copy number calculator converts mass to molecules. Browse the full free online tools hub for more.
Frequently Asked Questions
Add the average residue mass of every amino acid in the sequence, then add 18.01524 daltons for the water molecule that completes the two free termini. Residue masses are the amino acid masses already less the water lost when each peptide bond forms.
Because forming each peptide bond releases one water. Working in residue masses removes that water from every unit, so a single water has to be added back at the end to account for the free amino and carboxyl termini.
Average mass, which uses natural isotopic abundances and is the correct figure for converting between grams and moles. Monoisotopic mass, used for matching high-resolution mass spectrometry peaks, is lower and diverges further as the protein gets larger.
Tyrosines multiplied by 1,490, plus tryptophans multiplied by 5,500, plus cystines multiplied by 125, which is the method set out in the ExPASy ProtParam documentation. Free cysteines are excluded because they absorb negligibly at 280 nm.
SDS-PAGE separates by detergent binding rather than mass, and unusually acidic sequences, proline-rich regions and post-translational modifications all distort mobility. Glycosylation in particular can add tens of kilodaltons to the apparent size.
Yes, if the protein you are working with still has it. A hexahistidine tag, linker and cleavage site are real mass, and a tryptophan-rich fusion partner changes the extinction coefficient enough to skew any concentration measurement.
No. This tool parses amino acid single-letter code. For an inorganic compound written as a chemical formula, use a molar mass calculator instead, which parses element symbols and subscripts.
This calculator provides general educational estimates from the sequence you supply and does not account for post-translational modification, processing or bound ligands. Confirm masses experimentally before relying on them.
