Calculate protein molecular weight from amino acid sequence
Paste your protein sequence and the calculator identifies each amino acid residue, looks up its monoisotopic or average molecular weight, then sums them all. It subtracts (n-1) x 18.02 Da to account for the water molecules lost during peptide bond formation, giving you the total molecular weight of the intact polypeptide chain.
Knowing a protein's molecular weight is fundamental for interpreting SDS-PAGE gels, calibrating size-exclusion chromatography columns, and confirming identity by mass spectrometry. It also guides buffer preparation for techniques like Western blotting and helps researchers determine oligomeric states by comparing native and denatured molecular weights in analytical ultracentrifugation experiments.
Molecular weight is expressed in Daltons (Da), where one Dalton equals one-twelfth the mass of a carbon-12 atom. Proteins typically range from a few thousand to several hundred thousand Daltons, so kilodaltons (kDa) are more convenient. For example, a 50,000 Da protein is written as 50 kDa — a shorthand used universally in biochemistry publications and lab protocols.
Always use single-letter IUPAC amino acid codes and remove any FASTA headers, spaces, or numbering before pasting your sequence. Remember that post-translational modifications like glycosylation, phosphorylation, or disulfide bonds alter the actual mass observed experimentally. For recombinant proteins, include any tags (His-tag, GST) that will be present during your downstream analysis.
Proteins are polymers of amino acids linked by peptide bonds. The properties of a protein are determined by its amino acid composition and sequence.
The molecular weight of a protein is calculated by summing the molecular weights of all amino acid residues and adding the weight of water (18.015 Da) for the peptide bonds formed.
• Average residue weight: ~110 Da
• Free amino acid average: ~138 Da
• Water loss per peptide bond: 18 Da
The extinction coefficient at 280 nm is calculated based on the number of tryptophan (W), tyrosine (Y), and cystine (disulfide-bonded cysteine) residues:
ε₂₈₀ = (nW × 5500) + (nY × 1490) + (nCystine × 125) M⁻¹cm⁻¹
The isoelectric point is the pH at which the protein has no net charge. It depends on the relative numbers of acidic (Asp, Glu) and basic (Arg, Lys, His) amino acids.
| Class | Amino Acids | Properties |
|---|---|---|
| Nonpolar | A, V, L, I, M, F, W, P, G | Hydrophobic, interior of proteins |
| Polar uncharged | S, T, C, Y, N, Q | Hydrophilic, surface of proteins |
| Positively charged | K, R, H | Basic, often active sites |
| Negatively charged | D, E | Acidic, often active sites |
Protein molecular weight is calculated by summing the average molecular weight of each amino acid residue in the sequence, then adding 18.015 Da for the water molecule present at the termini. Each amino acid has a characteristic residue weight (the weight after losing water during peptide bond formation), ranging from 57.05 Da for glycine to 186.21 Da for tryptophan. The average residue weight is approximately 110 Da, so a quick estimate is sequence length multiplied by 110.
A Dalton (Da) is the standard unit of molecular mass used in biochemistry, defined as one-twelfth the mass of a carbon-12 atom (approximately 1.66 x 10^-24 grams). It is numerically equivalent to grams per mole (g/mol). Proteins are often measured in kilodaltons (kDa), where 1 kDa equals 1,000 Da. For example, a typical antibody has a molecular weight of about 150 kDa, while insulin is approximately 5.8 kDa.
Several factors cause discrepancies between calculated and apparent molecular weight on SDS-PAGE. Post-translational modifications like glycosylation add mass that increases apparent size. Highly charged or hydrophobic proteins may bind SDS differently, altering their migration. Proteins with high proline content or intrinsically disordered regions often appear 10 to 20 percent larger than expected. Membrane proteins and very basic proteins are also notorious for anomalous migration. Mass spectrometry provides the most accurate experimental molecular weight.