Drybench Sequence product page
Drybench Sequence.
GFP: ε280, MW and pI with a His tag (example gfp@1)
A versioned teaching example: green fluorescent protein P42212 with and without a His6 tag, by the methods ProtParam uses.
Sequence
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Answer
ε280
The molar extinction coefficient at 280 nm by Pace et al.
More about this
1995, with all cysteines paired and all reduced, and the absorbance of a 1 g/L solution.
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Worked example
- Aequorea victoria green fluorescent protein, UniProt P42212, 238 residues: ε280 in both cysteine conventions, MW and pI.
- ε280 22,015 M⁻¹cm⁻¹ with the cysteines paired and 21,890 reduced; 26886.32 Da, pI 5.67.
- With a His6 tag and a TEV site in front (20 residues) the mass is 29154.69 Da and the pI 5.94; the ε280 does not change, since the tag has no Trp or Tyr. The protein page finds the tag and proposes the cut.
- Source: UniProtKB P42212, The UniProt Consortium (CC BY 4.0).
This page is the example gfp@1: its method versions are pinned, so it gives the same numbers next year. seq 1.1.0, 2026-10-06.
Method
ε280 = 5,500·nTrp + 1,490·nTyr + 125·n(cystine); A280(1 g/L) = ε280 / MW
- Molar extinction coefficient at 280 nm. Pace CN, Vajdos F, Fee L, Grimsley G, Gray T. How to measure and predict the molar absorption coefficient of a protein. Protein Sci 1995;4:2411-2423 (read 5 October 2026).Checked against Expasy ProtParam exactly on 22 sequences in both cysteine conventions (6 October 2026). ProtParam is a tool of the SIB Swiss Institute of Bioinformatics; not affiliated.
- Average molecular weight from the sequence. ExPASy ProtParam (Gasteiger et al. 2005); the fixtures are ProtParam's printed weights, retrieved 5 October 2026.Checked against Expasy ProtParam within 0.01 Da on 22 sequences with ProtParam's residue masses (6 October 2026). ProtParam is a tool of the SIB Swiss Institute of Bioinformatics; not affiliated.
- Theoretical isoelectric point. Bjellqvist B et al. Electrophoresis 1993;14:1023-1031; 1994;15:529-539; as ExPASy ProtParam applies them.Checked against Expasy ProtParam within 0.01 on 22 sequences, never a bound of the pH 0 to 14 search (6 October 2026). ProtParam is a tool of the SIB Swiss Institute of Bioinformatics; not affiliated.
- Tags and protease sites by motif. Peptide tag and protease recognition motifs from the cited papers; a cut is proposed at the site nearest an end and confirmed by the person, never inferred from the fold.Unit-tested on the fixture construct: the His6-TEV construct's cleavage product is the untagged fixture sequence (6 October 2026).
Constants
- pace-1995: Pace et al. 1995 coefficients: 5,500 Trp, 1,490 Tyr, 125 cystine
- protparam: ExPASy ProtParam average residue masses
- bjellqvist-protparam: Bjellqvist pK values as ExPASy ProtParam applies them, including a C-terminal pK of 3.55
Limits
- For research and education only, and not medical advice: nothing here is an assessment of any person's variant, sample or molecule, and a difference reported from a reference is a difference, not an interpretation.
- Hairpin and dimer ΔG are two-state estimates at 37 °C from the SantaLucia and Hicks 2004 rules without the special triloop and tetraloop bonuses, so hairpins with GNA or GNRA loops are estimated less stable than they are.