Peptide Molecular Weight Calculator

How to use the peptide molecular weight calculator

The calculator reads a sequence in one-letter code and applies the same bookkeeping a chemist does by hand. It updates as you type and stores your inputs in the page link.

1. Paste the sequence. Use one-letter codes from the N-terminus to the C-terminus, for example DRVYIHPF. Spaces, line numbers and a FASTA header line are stripped. Lowercase is fine, and so is hyphenated three-letter code such as Gly-His-Lys. If you write Ac- or -NH₂ at the ends, the terminal settings switch to match.
2. Pick the N-terminus. A free amine is the default. Choose acetylated if the sequence is written Ac-…
3. Pick the C-terminus. A free acid (…-OH) is the default. Choose amide if the sequence ends in -NH₂, as many peptide hormones do.
4. Enter disulfide bonds. Count Cys–Cys bridges, not cysteines. Two cysteines make one bond.
5. Read the results. The large number is the average molecular weight. Below it are the monoisotopic mass, the molecular formula, [M+H]⁺ to [M+3H]³⁺, the extinction coefficient and a composition table. Open Show the math to see each step.

How peptide molecular weight is calculated

A peptide is a chain of amino acids joined by peptide bonds. Inside the chain each amino acid is called a residue: the free amino acid minus one water. The molecular weight of the chain is the sum of its residues, plus the one water that remains at the two ends, plus or minus any modifications.

Every residue mass on this page is computed from its elemental formula. Monoisotopic masses use the NIST masses of ¹²C, ¹H, ¹⁴N, ¹⁶O and ³²S. Average masses use the IUPAC standard atomic weights listed by Unimod (C 12.0107, H 1.00794, N 14.0067, O 15.9994, S 32.065). ExPASy tools use slightly older atomic weights, so their averages run about 0.01–0.03 Da higher for a 1–3 kDa peptide. Monoisotopic values agree to the fourth decimal.

Why you add one water: the peptide bond

A peptide bond forms when the carboxyl group of one amino acid reacts with the amine of the next. The OH from the acid and one H from the amine leave together as water. This is a condensation reaction. A chain of n amino acids has n − 1 peptide bonds and has lost n − 1 waters, which is why the formula uses residue masses and adds back exactly one water.

Amino acid molecular weight table

The table lists all 20 standard amino acids. The residue columns are what you add up for a peptide. The last column is the free amino acid (residue + H₂O), which is the number to use when you weigh out a single amino acid. Each value is computed from the formula in the third column, so the table and the calculator can never disagree.

Leucine and isoleucine share the formula C₆H₁₁NO, so no mass measurement can tell them apart. Lysine and glutamine differ by only 0.0364 Da monoisotopic, which takes high resolution to separate.

Average vs monoisotopic mass

Carbon is about 98.9% ¹²C and 1.1% ¹³C. Hydrogen, nitrogen, oxygen and sulfur also have heavier isotopes. A real sample is therefore a mix of molecules that differ by whole neutron steps. Two numbers describe it.

• Monoisotopic mass uses only the lightest isotope of each element. It is the first peak of the isotope cluster in a high-resolution mass spectrum.
• Average molecular weight uses the standard atomic weights, which blend every isotope by natural abundance. It is the right number for weighing, molarity and anything done on a balance.

The gap between the two numbers grows with size, because every extra carbon adds another chance of a ¹³C. Around 2 kDa the M+1 peak overtakes the monoisotopic peak, and for large proteins the monoisotopic peak can almost disappear. The table shows the trend, with the tallest peak computed from each formula.

Masses in Da. All sequences have free termini unless noted.

Modifications and terminal groups

Many research and therapeutic peptides are not bare chains. A modification changes the elemental formula, and the mass change follows directly from it. The deltas below are computed from Unimod compositions with the same atomic masses as the calculator.

The disulfide row is per bond (two Cys). Unimod lists it per cysteine as “Dehydro”, −H.

Two modifications change charge as well as mass. Acetylation removes the positive N-terminus and amidation removes the negative C-terminus, so both shift the isoelectric point. Phosphorylation and deamidation add negative charge. Run the modified form through the isoelectric point calculator to see the effect.

Mass spec m/z values for [M+nH]ⁿ⁺

A mass spectrometer measures mass-to-charge ratio (m/z), not mass. In positive mode, peptides pick up protons. Electrospray often gives several charge states at once, while MALDI mostly gives the singly charged [M+H]⁺. Each added proton adds 1.00728 Da and one charge. Use the monoisotopic mass for high-resolution data.

To check a charge state in your spectrum, look at the isotope spacing. Peaks of a 2+ ion sit about 0.5 m/z apart, 3+ about 0.33 apart. Sodium and potassium adducts appear as extra peaks above [M+H]⁺, and oxidized methionine adds 16 Da per site.

Worked examples

Example 1: angiotensin II, DRVYIHPF

The eight residues sum to 1,028.1634 Da. Adding 18.0153 Da of water gives 1,046.18 Da, formula C₅₀H₇₁N₁₃O₁₂. The monoisotopic mass is 1,045.5345 Da, so [M+H]⁺ is 1046.5418 and [M+2H]²⁺ is 523.7745. PubChem lists the same formula and a molecular weight of 1046.2.

Example 2: BPC-157, GEPPPGKPADDAGLV

Fifteen residues sum to 1,401.5202 Da. With water the average molecular weight is 1,419.54 Da and the monoisotopic mass is 1,418.7042 Da. BPC-157 is an investigational peptide, not an FDA-approved drug; the BPC-157 calculator covers its status and vial math.

Example 3: oxytocin with an amide and a disulfide

Oxytocin is CYIQNCPLG with a C-terminal amide and one disulfide between its two cysteines. The free-acid, reduced chain is 1,010.19 Da. Amidation takes it to 1,009.20 Da. The disulfide removes two hydrogens, giving 1,007.19 Da, formula C₄₃H₆₆N₁₂O₁₂S₂. Skipping either correction would put the answer 3.00 Da too high.

Example 4: from milligrams to micromoles

Molecular weight turns a weighed amount into moles. 5 mg of BPC-157 is 5 mg ÷ 1,419.54 g/mol = 3.522 µmol. Dissolved in 2 mL, the solution is 1.761 mM. If your amount is written in micrograms, convert it with the mcg to mg converter first. For the volume-per-dose side of the math, use the peptide reconstitution calculator.

Extinction coefficient and A280

Tryptophan and tyrosine absorb ultraviolet light at 280 nm, and so do disulfide bonds, weakly. Pace and colleagues (1995) showed that a protein's molar extinction coefficient (ε) can be predicted from its composition. ExPASy ProtParam uses the same values.

Angiotensin II has one tyrosine, so ε is 1,490 M⁻¹cm⁻¹ and a 1 mg/mL solution reads about 1.424 in a 1 cm cuvette. Oxytocin adds one cystine to its tyrosine for 1,615. BPC-157 has no Trp or Tyr, so it barely absorbs at 280 nm; the calculator warns you when that happens.

Molecular weight vs net peptide content

Molecular weight describes one molecule. A vial of lyophilised peptide is a powder that can also contain counter-ions, water and residual salts. Peptides purified by reversed-phase HPLC with trifluoroacetic acid are often supplied as trifluoroacetate (TFA) salts, with counter-ions paired to positively charged groups. The share of the powder that is actually peptide is called the net peptide content. When a certificate of analysis gives it, multiply the gross weight by it before you convert to moles. For dosing arithmetic based on a labelled amount, the reconstitution calculator and the peptide blend calculator work from the label directly.

Common mistakes

• Letters that are not amino acids. B, J, O, U, X and Z are ambiguous or nonstandard codes. The calculator ignores them and shows a red warning, so check that the remaining length matches your peptide.
• More disulfides than cysteine pairs. Each bond needs two Cys. If you enter more bonds than the sequence allows, only the possible number is counted and you are warned.
• Three-letter code in capitals without hyphens. Gly-His-Lys and GlyHisLys are read as three residues, but GLYHISLYS is read as nine one-letter residues. An empty box gives no result at all.
• Forgetting the ends. An amidated or acetylated peptide entered as a free chain is off by about 1 or 42 Da, which is enough to miss the right peak.
• Adding free amino acid masses. Summing free amino acids instead of residues overcounts one water per peptide bond.
• Comparing the wrong mass. Match high-resolution spectra to the monoisotopic mass and weighing or SDS-PAGE estimates to the average.
• Trusting A280 without Trp or Tyr. If ε is zero or small, measure concentration another way.

Key terms

Residue

An amino acid inside a chain, equal to the free amino acid minus one water.

Peptide bond

The amide bond between the carboxyl carbon of one residue and the nitrogen of the next.

Dalton (Da)

The unified atomic mass unit. 1 Da per molecule corresponds to 1 g/mol.

Average molecular weight

Mass computed with standard atomic weights, which include all natural isotopes.

Monoisotopic mass

Mass computed with only the lightest isotope of each element.

m/z

Mass-to-charge ratio, the quantity a mass spectrometer measures.

Extinction coefficient (ε)

How strongly one mole per litre absorbs light over a 1 cm path at a given wavelength.

Net peptide content

The fraction of a lyophilised powder, by weight, that is peptide rather than counter-ions, water or salts.

Frequently asked questions

Sources

We check every formula and figure on this page against primary sources. See our editorial policy for how we review content.