peff-digest 0.1.0

PEFF-aware in-silico protein digestion with PTM enumeration, sequence variants, and ProForma output.

peff_digest

A PEFF-aware protein digest tool. Given a PEFF file and enzymatic digestion parameters, produces a CSV of peptides with their sequence (ProForma notation), variant annotation, length, and monoisotopic mass.

Each PEFF VariantSimple and VariantComplex annotation is applied independently (not combined). PEFF PTMs (ModResPsi / ModResUnimod) are applied combinatorially up to a configurable limit per peptide. Fixed and variable user-defined modifications — including terminal modifications — are also supported.

Installation

uv sync
# or
just install

Requires Python 3.12+.

Usage

Via config file

peff-digest --config config.toml

Via flags

peff-digest --peff-file human.peff --output-file peptides.csv

Flags override any values set in --config. Run peff-digest --help for the full flag reference.

Output

The output CSV has five columns:

Column	Description
protein_id	db_unique_id from the PEFF header
sequence	ProForma-annotated peptide sequence (includes mods)
variant	PEFF variant notation, e.g. (42|R), or empty for canonical
length	Peptide length in residues
mass	Monoisotopic mass in Da, or empty if not computable

Config reference

All options can be set in a TOML or JSON config file. TOML example:

peff_file = "human.peff"
output_file = "peptides.csv"

cleave_on = "KR"
missed_cleavages = 2
semi_enzymatic = false
max_ptm_per_peptide = 2
min_length = 7
max_length = 40
restrict_after = "P"
restrict_before = ""
cterminal = true
min_mass = 400.0
max_mass = 10000.0
drop_invalid_mass = false
annotate_variants = true

# Internal modifications — one [[internal_mods]] block per mod:
# [[internal_mods]]
# modification = "Carbamidomethyl"
# residue = "C"
# mod_type = "fixed"
#
# [[internal_mods]]
# modification = "Oxidation"
# residue = "M"
# mod_type = "variable"

# Terminal modifications — one [[terminal_mods]] block per mod:
# [[terminal_mods]]
# modification = "Acetyl"
# position = "nterm"
# mod_type = "variable"
# protein_terminus = true   # only the first peptide of each protein
#
# [[terminal_mods]]
# modification = "UNIMOD:737"
# position = "nterm"
# mod_type = "fixed"
# residue = "M"             # only if the terminal residue is M
#
# [[terminal_mods]]
# modification = "Amidated"
# position = "cterm"
# mod_type = "variable"

DigestConfig fields

Field	Type	Default	Description
peff_file	str	required	Path to the input PEFF file. Must exist.
output_file	str	"peptides.csv"	Path for the output CSV.
cleave_on	str	"KR"	Amino acids at which to cleave (e.g. "KR" for trypsin).
missed_cleavages	int	2	Maximum number of missed cleavage sites per peptide. Min 0.
semi_enzymatic	bool	false	Include semi-enzymatic peptides (one non-enzymatic terminus).
max_ptm_per_peptide	int	2	Maximum number of variable mods (PEFF + user) applied simultaneously per peptide. 0 disables all variable mods. Min 0.
min_length	int	7	Minimum peptide length in residues (inclusive). Min 1.
max_length	int	40	Maximum peptide length in residues (inclusive). Min 1.
restrict_after	str	"P"	Skip cleavage when the following residue is in this set (e.g. "P" for trypsin/Pro rule).
restrict_before	str	""	Skip cleavage when the preceding residue is in this set.
cterminal	bool	true	true = C-terminal cleavage (standard); false = N-terminal.
internal_mods	list[InternalMod]	[]	Per-residue modifications. See InternalMod fields below.
terminal_mods	list[TerminalMod]	[]	Terminal modifications. See TerminalMod fields below.
min_mass	float | None	None	Minimum peptide mass in Da. Ignored if None.
max_mass	float | None	None	Maximum peptide mass in Da. Ignored if None.
drop_invalid_mass	bool	false	If true, exclude peptides whose mass cannot be computed.
annotate_variants	bool	true	If false, do not set peptide_name on variant peptides.
workers	int | None	None	Number of worker processes. Defaults to all available CPUs. Min 1.

InternalMod fields

Field	Type	Default	Description
modification	str	required	Modification name (e.g. "Carbamidomethyl", "UNIMOD:21").
residue	str	required	One or more amino acids the mod applies to (e.g. "C" or "KR").
mod_type	"fixed" | "variable"	required	"fixed" = always applied; "variable" = enumerated combinatorially (counts against max_ptm_per_peptide).

TerminalMod fields

Field	Type	Default	Description
modification	str	required	Modification name (e.g. "Acetyl", "UNIMOD:737").
position	"nterm" | "cterm"	required	Which terminus to apply the mod to.
mod_type	"fixed" | "variable"	required	"fixed" = always applied; "variable" = enumerated combinatorially (counts against max_ptm_per_peptide).
residue	str | None	None	If set, the mod is only applied when the terminal residue is in this string (e.g. "M" or "KR").
protein_terminus	bool	false	If true, only apply to the protein-level terminus (first peptide for N-term, last for C-term).

Python API

Full digest → Polars DataFrame

from peff_digest import DigestConfig, InternalMod, TerminalMod, digest

config = DigestConfig(
    peff_file="human.peff",
    missed_cleavages=2,
    min_length=7,
    max_length=40,
    min_mass=400.0,
    max_mass=10000.0,
    drop_invalid_mass=True,
    internal_mods=[
        InternalMod(modification="Carbamidomethyl", residue="C", mod_type="fixed"),
        InternalMod(modification="Oxidation", residue="M", mod_type="variable"),
    ],
    terminal_mods=[
        TerminalMod(modification="Acetyl", position="nterm", mod_type="variable", protein_terminus=True),
    ],
)

df = digest(config)
print(df)

Returns a polars.DataFrame with columns protein_id, sequence, variant, length, mass. All filtering from the config (mass bounds, drop_invalid_mass) is applied before returning.

Single-entry digest

import pefftacular as pf
from peff_digest import InternalMod, TerminalMod, digest_peff_sequence

entry = next(iter(pf.PeffReader("human.peff")))

peptides = digest_peff_sequence(
    entry,
    cleave_on="KR",
    missed_cleavages=2,
    min_length=7,
    max_length=40,
    restrict_after="P",
    internal_mods=[
        InternalMod(modification="Carbamidomethyl", residue="C", mod_type="fixed"),
        InternalMod(modification="Oxidation", residue="M", mod_type="variable"),
    ],
    max_ptm_per_peptide=2,
    terminal_mods=[
        TerminalMod(modification="Acetyl", position="nterm", mod_type="variable", protein_terminus=True),
        TerminalMod(modification="Amidated", position="cterm", mod_type="variable"),
    ],
)

for peptide in peptides:
    print(str(peptide), len(peptide), peptide.mass())

Returns a set[peptacular.ProFormaAnnotation]. Each element supports len(), .mass(), str(), and .peptide_name (PEFF variant notation, or None for canonical).

Development

just lint      # ruff check
just format    # ruff format + import sort
just test      # pytest
just check     # lint + type check (ty) + test