Reading peptide sequence notation: one-letter, three-letter codes and modification symbols

Two alphabets for the same chain

The IUPAC-IUB nomenclature recommendations (JCBN) define both systems. Sequences are written N-terminus on the left, C-terminus on the right. [1] The one-letter code is designed for compact comparison of long sequences in tables and lists; the recommendations say it is harder to read than the three-letter code and should not be used in simple text or when reporting experimental sequence details. [1]

Supplier listings often use one-letter strings because they fit in a product title. That is fine for searching, but a one-letter string has no standard place for termini, D-residues or bridges. The EMA guideline for synthetic peptide medicines asks for three-letter codes per IUPAC, with a legend for any non-standard symbol and both termini clearly indicated. [6]

Reference table: the 20 standard amino acids

One- and three-letter codes are from IUPAC-IUB; residue masses (the mass each residue contributes inside a chain, without terminal water) are from ExPASy. [1] [5] Note the pairs with identical or near-identical masses: leucine and isoleucine are identical, and glutamine and lysine differ by only 0.036 Da.

Ambiguity codes and non-standard symbols

IUPAC also defines B (Asx, aspartic acid or asparagine when not distinguished), Z (Glx, glutamic acid or glutamine when not distinguished), X (Xaa, unknown or other) and U (Sec, selenocysteine). [1] A B, Z or X in a supplier sequence for a catalog peptide means the supplier is telling you it does not know which residue is there, or is using X as a placeholder for something non-standard. Either way, ask.

Within a written sequence, IUPAC uses a slash at an end to mean the fragment is not known to be the complete molecule and parentheses for a region of known composition but undetermined order. [1] Those conventions belong to sequencing reports, but if they appear on a product page they signal uncertainty, not shorthand.

Termini, D-residues and other marks

IUPAC states that residue symbols in a sequence denote the L configuration for chiral amino acids unless otherwise indicated, and that a D residue is shown by inserting D- before the symbol, as in Phe-D-Ala-Leu. [2] [3] So a sequence with no prefixes claims every chiral residue is L. If a catalog peptide is meant to contain a D residue and the listing drops the prefix, the listing describes a different stereoisomer.

Common terminal marks: H- for a free N-terminus and -OH for a free C-terminal acid (Bachem writes H-Gln-...-Gly-OH), -NH2 for an amide, Ac- for acetyl and Glp for pyroglutamyl, as in Glp-His-Pro-NH2. Cyclic peptides are written cyclo( ). [2] [7] Our guide to terminal modifications gives the mass change for each.

Worked check: a supplier string against UniProt

UniProt entry P01178 is the human oxytocin-neurophysin 1 precursor, 125 residues long. Oxytocin is recorded as residues 20 to 28, with a glycine amide at 28 and a disulfide between 20 and 25. [4] Reading positions 20 to 28 from the flat-file sequence gives CYIQNCPLG; in three-letter form with features, H-Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly-NH2 with a Cys1-Cys6 disulfide.

Two traps appear immediately. First, the precursor continues GKR after residue 28, and copying a few characters too many gives CYIQNCPLGGK, an 11-residue string. Second, the amide is a separate feature line, not part of the sequence string; a supplier who copies only the string loses it. Now take an invented supplier listing: "CYLQNCPLG, MW 1007.2". Aligned with UniProt, position 3 reads L instead of I. Leucine and isoleucine have identical residue masses, so the listed weight and any intact-mass result would still match oxytocin. [5] The EMA lists "Ile in Leu and vice versa" among impurities that come from starting materials, so this is not only a typing risk. [6]

Sequence comparison checklist

A string match tells you the supplier wrote the right letters, not what is in the vial. Pair it with a certificate that confirms identity by a method that can see the difference you care about: amino acid analysis for composition questions such as a Leu-for-Ile swap, and chiral analysis for D/L questions.

• Count residues in both strings before aligning.
• Align position by position; flag every mismatch, including same-mass pairs (I/L) and near-same pairs (Q/K).
• Check the termini: H- or Ac-, -OH or -NH2.
• Check D- prefixes; no prefix means L.
• Check disulfide pairings and cyclisation are stated.
• Check for B, Z or X and ask what they stand for.
• Record the database accession and position range you compared against.