33,061.52
Da
33.062
kDa
300
299
33,061.52
Da
33.062
kDa
300
299
The Protein Molecular Weight Calculator estimates the molecular weight of a protein based on its amino acid composition. Molecular weight (MW) is a fundamental property of proteins, critical for techniques such as SDS-PAGE gel interpretation, size-exclusion chromatography, mass spectrometry validation, and buffer preparation. This tool computes the sum of individual amino acid residue masses minus the water molecules lost during peptide bond formation.
The calculator supports two modes: a quick estimate using the total number of amino acids with an average residue mass (~128.16 Da), and a precise calculation using the exact count of each of the 20 standard amino acids. The average human protein contains approximately 375 residues, while the largest known protein, titin, exceeds 34,000 residues with a molecular weight above 3.8 MDa.
The molecular weight of a protein is calculated by summing the masses of all constituent amino acid residues and subtracting the water released during peptide bond formation:
$$MW = \sum_{i=1}^{n} MW_i - (n-1) \times 18.015$$
where $$MW_i$$ is the molecular weight of amino acid residue $$i$$, $$n$$ is the total number of residues, and $$18.015 \text{ Da}$$ is the molecular weight of water lost per peptide bond. Each peptide bond forms via a condensation reaction that releases one water molecule. For a protein with $$n$$ residues, there are $$(n-1)$$ peptide bonds.
When using the average residue method, the formula simplifies to:
$$MW \approx n \times \overline{MW}_{residue} - (n-1) \times 18.015$$
The average residue molecular weight of 128.16 Da reflects the weighted mean of all 20 standard amino acids based on typical proteome composition. Individual residue masses range from 75.03 Da (glycine) to 204.23 Da (tryptophan).
The calculated molecular weight represents the theoretical mass of the unmodified polypeptide chain. Post-translational modifications (PTMs) such as glycosylation, phosphorylation, ubiquitination, and disulfide bond formation will alter the actual observed mass. Glycosylation alone can add 1-80 kDa or more.
On SDS-PAGE, apparent molecular weight may differ from the calculated value by 5-15% due to anomalous migration caused by proline-rich regions, highly charged segments, or incomplete SDS binding. For mass spectrometry, the calculated MW provides a reference for intact protein analysis and peptide mass fingerprinting. Proteins below ~10 kDa are considered small peptides/proteins, 10-100 kDa covers most enzymes and structural proteins, while those above 100 kDa are large multidomain proteins.
Inputs
Results
Using the average amino acid MW of 128.16 Da for a 375-residue protein yields approximately 41.5 kDa, consistent with the median size of human proteins.
Inputs
Results
GFP contains 238 amino acids with a known MW of ~26.9 kDa. The average residue estimate gives ~26.4 kDa, within 2% of the actual value.
Peptide bond formation is a condensation reaction. When two amino acids join, the amino group of one reacts with the carboxyl group of another, releasing one water molecule (18.015 Da). For a protein with n residues, there are (n-1) peptide bonds, so (n-1) water molecules are lost from the total mass.
The value of 128.16 Da represents the weighted mean of all 20 standard amino acids based on average proteome composition. For most proteins, this gives estimates within 2-5% of the true mass. Proteins enriched in heavy residues (Trp, Tyr) will be underestimated, while glycine-rich proteins will be overestimated.
No. The calculator computes the mass of the unmodified polypeptide chain only. Common PTMs like phosphorylation (+80 Da), glycosylation (+162+ Da per sugar), acetylation (+42 Da), and methylation (+14 Da) must be added separately. For glycoproteins, the sugar moieties can add substantial mass.
A Dalton (Da) is the standard unit of atomic and molecular mass, approximately equal to the mass of one hydrogen atom (1.66 x 10^-24 g). A kiloDalton (kDa) equals 1,000 Da. Proteins are commonly expressed in kDa for convenience, with most falling in the 10-200 kDa range.
SDS-PAGE estimates MW based on electrophoretic mobility, which assumes uniform charge-to-mass ratio from SDS binding. Anomalous migration occurs with proline-rich proteins, highly charged proteins, membrane proteins with bound lipids, and glycoproteins. Differences of 10-20% are not uncommon for these protein types.
Amino acid composition can be obtained from: (1) the protein sequence from databases like UniProt, (2) amino acid analysis by acid hydrolysis, or (3) mass spectrometry-based sequencing. The ExPASy ProtParam tool can compute composition from a FASTA sequence for comparison.
Small peptides and hormones: 0.5-10 kDa. Enzymes and regulatory proteins: 10-80 kDa. Antibodies (IgG): ~150 kDa. Structural proteins (collagen): ~300 kDa. Muscle proteins (titin): ~3,800 kDa. Viral capsid proteins can form assemblies exceeding 10,000 kDa.
This calculator is designed for polypeptide chains only. For ribonucleoprotein complexes, calculate the protein and nucleic acid masses separately and sum them. Each nucleotide adds approximately 330 Da (DNA) or 340 Da (RNA) to the complex mass.
Each disulfide bond (S-S) between two cysteine residues results in the loss of 2 hydrogen atoms (2.016 Da). For a protein with d disulfide bonds, subtract d x 2.016 Da from the calculated MW. This correction is small but relevant for mass spectrometry applications.
Methionine has a residue MW of 149.21 Da. In many organisms, the N-terminal methionine is cleaved post-translationally by methionine aminopeptidase if the second residue is small (Ala, Cys, Gly, Pro, Ser, Thr, Val). This removes ~131 Da (residue MW minus water) from the final protein mass.
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