The difference between a peptide ending in -OH and one ending in -NH2 can be easy to overlook in a sequence label. It identifies a change at the terminal carboxyl group. The amino-acid order can remain the same while the terminal chemistry, calculated mass and potential interactions differ.
Read the group attached to the last carbonyl
For the common primary C-terminal amide, the terminal group is -C(=O)NH2 rather than -C(=O)OH. This is separate from the amide groups already present in the peptide backbone and from the side-chain amides of asparagine or glutamine.Unimod 2 — Amidation (opens in a new tab)
| Notation | Interpretation |
|---|---|
| Ala-Gly-OH | Free C-terminal carboxylic acid form |
| Ala-Gly-NH2 | C-terminal primary carboxamide |
| A sequence containing Asn | A side-chain amide is present; the terminus still needs its own annotation |
The word amidated should therefore be read with its location. Counting every amide in the molecular structure would answer a different question from identifying whether the C-terminus has been modified.
A terminal amide also should not be confused with adding ammonia as an unbound ingredient. The notation describes atoms connected within the peptide molecule.
A smaller mass is consistent with adding nitrogen
Unimod gives the net composition change as H + N − O and the monoisotopic mass change as −0.984016 Da. Although nitrogen is introduced, oxygen is replaced; the complete atom bookkeeping explains the negative sign.Unimod 2 — Amidation (opens in a new tab)
For an invented free-acid neutral mass of 1,000.000000 Da, the corresponding primary-amide calculation is 999.015984 Da. Comparing rounded integer masses would obscure much of this difference. The example uses neutral monoisotopic masses, not measured ion peaks.
The reverse change has the opposite sign when the comparison truly involves those same terminal groups. A generic deamidation annotation elsewhere in a sequence is not automatically evidence that the C-terminal amide changed; the site must be specified.
Terminal chemistry can matter without a universal rule
Kang and colleagues examined terminal amidation alongside loop-residue changes in conotoxin variants. Their results linked those chemical features to different disulfide-folding preferences in the studied systems. The finding is about defined sequences and comparisons.Kang and colleagues — Terminal chemistry and folding preferences (opens in a new tab)
It does not mean that every amidated peptide adopts one fold or resists every form of degradation. A terminal modification changes one part of the molecule; its practical consequence depends on the interactions and process being measured.
Likewise, removing a free terminal carboxyl group from a charge model does not establish the whole peptide's net charge. Acidic and basic side chains and the other terminus must still be considered under the stated conditions.
Keep the end group in every comparison
A parent peptide and an amidated analogue may share an informal name in a figure. Check the methods or structure legend before combining their results. A change that is omitted from a short name can still be central to the experiment.
For a clear record, preserve both terminal annotations with the sequence, including whether the N-terminus is free or capped. This prevents a double-terminal modification from being mistaken for a single C-terminal change.
- Identify the terminal carbonyl and its attached group.
- Keep C-terminal amidation separate from Asn/Gln side-chain chemistry.
- Use the appropriate reference structure for mass comparisons.
- Record measured effects only for the constructs actually studied.
Sources and further detail
- Unimod 2 — Amidation (opens in a new tab)
Elemental difference H + N − O and monoisotopic mass change −0.984016 Da for the specified modification.
- Kang and colleagues — Terminal chemistry and folding preferences (opens in a new tab)
Biochemistry 46, 3338–3355 (2007), DOI 10.1021/bi061969o. Conotoxin variant study examines terminal amidation and loop-residue changes; no universal performance rule is inferred.
Sources checked 19 September 2026. Worked examples are illustrative unless a supplied report is explicitly identified. This article has not undergone independent scientific peer review.