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Supplementary Dataset 1 - E coli proteome-derived peptide libraries made with trypsin, chymotrypsin and GluC

ABSTRACT: E. coli proteome-derived peptide libraries produced with trypsin, chymotrypsin, or GluC

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Escherichia Coli (ncbitaxon:562)

SUBMITTER: Amy Weeks

PROVIDER: MSV000091247 | MassIVE | Sat Feb 11 13:14:00 GMT 2023

SECONDARY ACCESSION(S): PXD040044

REPOSITORIES: MassIVE

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Supplementary Dataset 15 - Human proteome-derived peptide libraries generated with trypsin, chymotrypsin, and GluC

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Supplementary Dataset 20 - GluC PICS2 with E coli proteome-derived tryptic peptide library

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Project description:High specificity and ease of use make trypsin the most used enzyme in proteomics. Proteases with complementary cleavage specificity to trypsin have been applied to obtain additional data. However, use of proteases with broad specificity proved especially challenging. In this work, we analyzed the characteristics of five protease alternatives to trypsin for protein identification and sequence coverage when applied to S. pombe whole cell lysates. The specificity of the protease heavily impacted on the number of proteins identified. Proteases with higher specificity let to the identification of more proteins than proteases with lower specificity. However, AspN, GluC, chymotrypsin and proteinase K largely benefited from being paired with trypsin in sequential digestion, as had been shown by us for elastase before. In the most extreme case, the addition of trypsin to a proteinase K digest increased the number of identified proteins by 524 %. Also, AspN (82 %) and GluC (74 %) protein identifications largely improved following the additional digestion with trypsin. In general, protein identifications improved most over the use of the single protease when the enzymes followed on an initial digestion with trypsin. In the most extreme case, the sequential digest with trypsin and AspN yielded even higher number of protein identifications than digesting with trypsin alone.

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Inactivation of mesotrypsin by chymotrypsin C prevents trypsin inhibitor degradation.

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| S-EPMC7076226 | biostudies-literature