Protein Molecular Weight Calculator
Calculate the molecular weight of a protein from its amino acid sequence.
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What it does
The Protein Molecular Weight Calculator turns an amino acid sequence into its molecular mass in daltons, reported alongside the residue count and the average residue mass. The sequence can be pasted as single-letter codes such as MKWVTFISLL, as three-letter codes such as Met-Lys-Trp, or as a three-letter run with no separators such as MetLysTrp, and the tool states how it read the input so the interpretation is never hidden. Masses can be reported either as average mass or as monoisotopic mass. It is the tool for checking a calculated mass against an SDS-PAGE band, for estimating what a construct will contribute to a total protein reading, and for confirming that a synthetic peptide has the mass it should.
How it works
The mass of a peptide is the sum of its residue masses plus the mass of one water molecule. Residue masses are used rather than free amino acid masses because a residue is what remains after a peptide bond has formed and one water has been lost per bond, and the single water added back accounts for the free amino and carboxyl ends of the chain. Two conventions are available. Average residue masses weight each element by its natural isotopic abundance, giving 71.0788 for alanine and 18.0153 for the terminal water; these are the numbers to compare against a gel or a colorimetric assay. Monoisotopic residue masses take only the lightest stable isotope of each element, giving 71.03711 for alanine and 18.010565 for the water, and these are what a mass spectrometer reports. The gap is roughly 0.02 to 0.13 Da per residue and accumulates with length, so it is well under a dalton for a short peptide and several daltons for a protein. The average residue mass shown alongside the total is the molecular weight minus the water, divided by the number of residues, and for most proteins it sits close to 110 Da, which makes it possible to estimate a mass from a sequence length in a few seconds. No post-translational modification, disulfide bond or terminal methionine processing is included.
Worked example
Met-enkephalin is the pentapeptide YGGFM. Summing the residue masses gives 163.1760 + 57.0519 + 57.0519 + 147.1766 + 131.1926 = 555.6489, and adding the terminal water gives 555.6489 + 18.0153 = 573.66 Da, with an average residue mass of 111.13 Da. Switching to monoisotopic masses gives 573.23 Da for the same peptide, a gap of 0.43 Da, and by ten residues the gap has grown: MKWVTFISLL is 1237.57 Da as an average mass against 1236.69 Da monoisotopic. That is the rule to carry forward, use the average mass for a gel and the monoisotopic mass for a spectrum. The A chain of insulin, GIVEQCCTSICSLYQLENYCN, is 21 residues and comes to 2383.71 Da, an average residue mass of 112.65 Da. As a rule of thumb, multiply the residue count by about 110: a 300 residue protein is roughly 33 kDa, which is why a band that runs a long way from its predicted position deserves investigation. Modifications push in the other direction and are not counted here: each phosphorylation adds about 79.97 Da and each disulfide bond removes 2.02 Da, so a modified peptide weighs more than the sequence alone predicts while a cross-linked one weighs less.
When to use it
The tool always states how it read the input, because single-letter and three-letter codes can look alike. Separated sequences are unambiguous: Met-Lys-Trp, Met Lys Trp and MetLysTrp all read as three-letter codes, while MKWVTFISLL and M K W read as single letters. With no separators the parser first tries to cut the whole run into three-letter names, which is why AlaGlyTrp is recognised as the tripeptide AGW at 332.36 Da rather than read letter by letter as nine residues at 961.13 Da, and it falls back to single letters only when that split fails, which is why WATER is read as five residues and MKW as three. One case is genuinely ambiguous, a run of single letters that also splits into valid names such as ALAGLY, which is read as the dipeptide Ala-Gly; the format selector forces the reading you intended and the line under the input always states which one is in effect, so check it whenever the residue count looks wrong. Two more points before trusting a number: a sequence containing typos returns a lower and entirely believable mass, so compare the residue count against the warning list, and the value shown is an average mass unless monoisotopic is selected, which matters when checking a peptide against a mass spectrum.
FAQ
- How is the molecular weight of a peptide calculated?
- Add up the residue masses of every amino acid in the chain and add 18.0153 Da for the terminal water. YGGFM sums to 555.6489 as residues and 573.66 Da as the peptide. Residue masses are used because forming each peptide bond removes one water, and the single water added back restores the free amino and carboxyl ends.
- Why does my calculated mass differ from the mass spectrum?
- The difference is usually isotopic. This tool reports the average mass, built from natural-abundance atomic weights and dominated by carbon-13 and nitrogen-15 contributions, while a mass spectrometer reports the monoisotopic mass of the most abundant light isotopologue. Average masses run slightly higher and the gap widens with length. Check also for modifications the tool does not count, since phosphorylation adds 79.97 Da and each disulfide bond removes 2.02 Da, and for N-terminal methionine removal, which takes off 131.19 Da.
- How does the tool decide between single-letter and three-letter input?
- Separators decide first: with hyphens or spaces each token is judged on its own, so Met-Lys-Trp is three-letter and M K W is three single letters. With no separators it tries to cut the whole run into three-letter names, which is how AlaGlyTrp is recognised as the tripeptide AGW, and falls back to single letters when that fails, which is how WATER and MKWVTFISLL are read. The interpretation is printed under the input box, and the format selector overrides it when a run such as ALAGLY is genuinely ambiguous.
- Can I paste a sequence with spaces, numbers or ambiguous codes?
- Separators are welcome in three-letter form, and a single-letter sequence may be written with or without spaces. Digits, punctuation other than the hyphen or space, and any letter outside the twenty codes are listed as invalid and skipped, while the mass is still computed from the residues that were recognised. A sequence containing typos therefore returns a lower, and entirely believable, number, so compare the residue count against the warning list before using the result. Ambiguous codes such as B, Z or X are not amino acids and are skipped as well. Note that the masses are average masses unless monoisotopic is selected, because an average mass and a monoisotopic mass differ by more than a dalton for anything longer than a few residues.