Digestion

Proteolytic Digestion with Trypsin

OpenMS has classes for proteolytic digestion which can be used as follows:

import pyopenms as oms
from urllib.request import urlretrieve

gh = "https://raw.githubusercontent.com/OpenMS/pyopenms-docs/master"
urlretrieve(gh + "/src/data/P02769.fasta", "bsa.fasta")

dig = oms.ProteaseDigestion()
dig.getEnzymeName()  # Trypsin
bsa = "".join([l.strip() for l in open("bsa.fasta").readlines()[1:]])
bsa = oms.AASequence.fromString(bsa)
# create all digestion products
result = []
dig.digest(bsa, result)
print(result[4].toString())
len(result)  # 82 peptides

Very short peptides or even single amino acid digestion products are often discarded as they usually contain little information (e.g., can’t be used to identify proteins). We now only generate digestion products with a length of \(7\) to \(40\).

# only create peptides of length 7-40
dig.digest(bsa, result, 7, 40)
# print the results
for s in result:
    print(s.toString())

Enzymatic digestion is often not perfect and sometimes enzymes miss cutting a peptide. We now allow up to two missed cleavages.

# Allow two missed cleavages
dig.setMissedCleavages(2)
# only create peptides of length 7-40
dig.digest(bsa, result, 7, 40)
# print the results
for s in result:
    print(s.toString())

Proteolytic Digestion with Lys-C

We can of course also use different enzymes, these are defined in the Enzymes.xml file and can be accessed using the EnzymesDB object

names = []
oms.ProteaseDB().getAllNames(names)
len(names)  # at least 25 by default
e = oms.ProteaseDB().getEnzyme("Lys-C")
e.getRegExDescription()
e.getRegEx()

Now that we have learned about the other enzymes available, we can use it to cut our protein of interest:

from urllib.request import urlretrieve

gh = "https://raw.githubusercontent.com/OpenMS/pyopenms-docs/master"
urlretrieve(gh + "/src/data/P02769.fasta", "bsa.fasta")

dig = oms.ProteaseDigestion()
dig.setEnzyme("Lys-C")
bsa = "".join([l.strip() for l in open("bsa.fasta").readlines()[1:]])
bsa = oms.AASequence.fromString(bsa)
result = []
dig.digest(bsa, result)
print(result[4].toString())
len(result)  # 57 peptides

We now get different digested peptides (\(57\) vs \(82\)) and the fourth peptide is now GLVLIAFSQYLQQCPFDEHVK instead of DTHK as with Trypsin (see above).

Oligonucleotide Digestion

There are multiple cleavage enzymes available for oligonucleotides, these are defined Enzymes_RNA.xml file and can be accessed using the RNaseDB object

db = oms.RNaseDB()
names = []
db.getAllNames(names)
names
# Will print out all available enzymes:
# ['RNase_U2', 'RNase_T1', 'RNase_H', 'unspecific cleavage', 'no cleavage', 'RNase_MC1', 'RNase_A', 'cusativin']
e = db.getEnzyme("RNase_T1")
e.getRegEx()
e.getThreePrimeGain()

We can now use it to cut an oligo:

oligo = oms.NASequence.fromString("pAUGUCGCAG")

dig = oms.RNaseDigestion()
dig.setEnzyme("RNase_T1")

result = []
dig.digest(oligo, result)
for fragment in result:
    print(fragment)

print("Looking closer at", result[0])
print(" Five Prime modification:", result[0].getFivePrimeMod().getCode())
print(" Three Prime modification:", result[0].getThreePrimeMod().getCode())
for ribo in result[0]:
    print(ribo.getCode(), ribo.getMonoMass(), ribo.isModified())