Peptide Molecular Weight Calculator
Calculate both average molecular weight and monoisotopic mass of peptides, including modified and C-terminal amidated sequences. This calculator supports FASTA format, terminal modifications (acetylation, amidation), isoelectric point (pI), net charge, GRAVY, and mass-to-molar conversions.
This calculator supports both average molecular weight and monoisotopic mass calculations. Average masses use the UWPR mass table; monoisotopic masses use NIST atomic masses (¹²C, ¹H, ¹⁴N, ¹⁶O, ³²S). pI and charge calculations use the IPC_peptide pKa set (Kozlowski, 2016). Results are theoretical approximations — always verify with experimental data.
Calculate Average and Monoisotopic Peptide Mass
Use the Average / Monoisotopic toggle in the result section to switch between two calculation modes. Average molecular weight uses naturally occurring isotope abundances and is standard for sample weighing and solution preparation. Monoisotopic mass uses the most abundant isotope of each element and corresponds to the M+0 peak observed in high-resolution mass spectrometry (LC-MS/MS, MALDI-TOF).
Both values are calculated from the same amino acid sequence and terminal modification settings. The mass/mole conversion section automatically updates based on the selected mode.
Monoisotopic Mass of a C-Terminal Amidated Peptide
C-terminal amidation replaces the terminal carboxyl group (–COOH) with an amide (–CONH₂), resulting in a mass shift of −0.9840 Da compared to the free carboxyl form. In monoisotopic terms, this corresponds to the loss of ¹⁶O (15.9949 Da) and gain of ¹⁴N¹H (15.0109 Da).
To calculate: select the C-terminal Amidation checkbox, switch to Monoisotopic mode, and the calculator applies the correction automatically. For a peptide "ACDEFGHIK" with C-terminal amidation, the monoisotopic mass is 1,017.4702 Da versus 1,018.4542 Da without amidation.
Average vs Monoisotopic Mass — What's the Difference?
Average mass reflects the weighted mean of all naturally occurring isotopes (e.g., carbon = 12.011 Da). This is the mass you measure on an analytical balance and the standard for peptide synthesis specifications, dilution calculations, and solution preparation.
Monoisotopic mass uses only the lightest abundant isotope of each element (¹²C = 12.000, ¹H = 1.00783, ¹⁴N = 14.003, ¹⁶O = 15.995, ³²S = 31.972). This value matches the first isotope peak (M+0) in a mass spectrum and is the standard for peptide identification in proteomics workflows (database search, de novo sequencing).
The difference between average and monoisotopic mass increases with peptide size — approximately 0.03% per 1,000 Da. For a 5 kDa peptide, the difference is typically 1.5–2.5 Da.
How Modifications Affect Peptide Mass
Terminal modifications change the peptide mass by altering the chemical groups at the N- or C-terminus. This calculator supports the two most common synthetic peptide modifications:
When both modifications are applied simultaneously, the net mass change is +41.053 Da (average) or +41.027 Da (monoisotopic). Both modifications also affect pI by removing terminal ionizable groups.
ℹ️ Frequently Asked Questions
How is peptide molecular weight calculated?
The molecular weight is computed by summing the residue masses of each amino acid in the sequence, then adding one water molecule (18.015 Da) to account for the free N- and C-termini.
MW = Σ(residue masses) + H₂O (18.015 Da)
Each residue mass equals the amino acid's molecular weight minus one water molecule lost during peptide bond formation. This calculator supports both average isotopic masses (for bench-side weighing) and monoisotopic masses (for mass spectrometry). Switch between modes using the toggle above the result.
What is the difference between average and monoisotopic mass?
Average mass uses isotope-weighted averages (bench weighing); monoisotopic mass uses the lightest abundant isotope (MS analysis). See "Average vs Monoisotopic Mass" above for full details and use case guidance.
How do I calculate monoisotopic mass for a C-terminal amidated peptide?
Select the C-terminal Amidation checkbox and switch to Monoisotopic mode. Amidation applies a −0.98402 Da correction. See "Monoisotopic Mass of a C-Terminal Amidated Peptide" above for the full formula and a worked example.
How do N-terminal acetylation and C-terminal amidation affect MW?
Acetylation adds +42.04 Da (avg) / +42.01 Da (mono). Amidation changes MW by −0.98 Da. Both also shift pI by removing terminal ionizable groups. See "How Modifications Affect Peptide Mass" above for detailed values and combined effects.
What is the isoelectric point (pI)?
The isoelectric point is the pH at which the peptide carries zero net charge. This calculator estimates pI using the Henderson-Hasselbalch equation and a binary search algorithm across pH 0–14.
At pH = pI → net charge ≈ 0
The pI depends on ionizable side chains (D, E, C, Y, H, K, R) as well as the free N- and C-terminal groups. Modifications like acetylation or amidation remove terminal ionizable groups and shift pI.
This calculator uses the IPC_peptide pKa set from Kozlowski (2016), optimized on 16,882 experimentally measured peptide isoelectric points. This set achieves a prediction RMSD of ~0.25 pH units — approximately 60% more accurate than traditional textbook values for peptide pI estimation.
How do I convert μg to nmol for a peptide?
Use the formula:
nmol = (μg × 1000) / MW
For example, for 10 μg of a peptide with MW = 2,500 Da:
nmol = (10 × 1000) / 2500 = 4.0 nmol
This calculator performs this conversion automatically in the Mass / Mole Conversion section.
What is GRAVY and how is it calculated?
GRAVY (Grand Average of Hydropathy) is the mean hydropathy value of all amino acids in the sequence, using the Kyte-Doolittle scale.
GRAVY = Σ(hydropathy values) / sequence length
Positive GRAVY → hydrophobic peptide (e.g., membrane-spanning regions).
Negative GRAVY → hydrophilic peptide (e.g., soluble proteins).
Scale ranges from −4.5 (Arg, most hydrophilic) to +4.5 (Ile, most hydrophobic).
Which pKa set does this calculator use?
This calculator uses the IPC_peptide pKa set from Kozlowski (2016), published in Biology Direct 11:55. This set was computationally optimized using a basin-hopping algorithm on 16,882 experimentally measured peptide isoelectric points from three independent datasets (Gauci, PHENYX, SEQUEST).
Compared to traditional textbook values (e.g., Lehninger), the IPC_peptide set reduces pI prediction error by approximately 60% for peptides (RMSD ~0.25 pH units).
The nine pKa values used:
NH₂ = 9.564 · COOH = 2.383
Asp = 3.887 · Glu = 4.317 · His = 6.018
Cys = 8.297 · Lys = 10.517 · Tyr = 10.071 · Arg = 12.503
Reference: Kozlowski LP (2016) IPC — Isoelectric Point Calculator. Biology Direct 11:55. doi:10.1186/s13062-016-0159-9