PXD021703

PXD021703 is an original dataset announced via ProteomeXchange.

Title	ProAlanase is an effective alternative to trypsin for proteomics applications and disulfide bond mapping_part3
Description	Trypsin is the protease of choice in bottom-up proteomics. However, its application can be limited by the amino acid composition of target proteins and the pH of the digestion solution. In this study we characterize ProAlanase, a protease from the fungus Aspergillus niger that cleaves primarily on the C-terminal side of proline and alanine residues. ProAlanase achieves high proteolytic activity and specificity when digestion is carried out at acidic pH (1.5) for relatively short (2 h) time periods. To elucidate the potential of ProAlanase in proteomics applications, we conducted a series of investigations comprising comparative multi-enzymatic profiling of a human cell line proteome, histone PTM analysis, ancient bone protein identification, phosphosite mapping and de novo sequencing of a proline-rich protein and disulfide bond mapping in monoclonal antibody. The results demonstrate that ProAlanase is highly suitable for proteomics analysis of the arginine- and lysine-rich histones, enabling high sequence coverage of multiple histone family members. It also facilitates an efficient digestion of bone collagen thanks to the cleavage at the C-terminus of hydroxyproline which is highly prevalent in collagen. This allows to identify complementary proteins in ProAlanase- and trypsin-digested ancient bone samples, as well as to increase sequence coverage of non-collagenous proteins. Moreover, digestion with ProAlanase improves protein sequence coverage and phosphosite localization for the proline-rich protein Notch3 intracellular domain (N3ICD). Furthermore, we achieve a nearly complete coverage of N3ICD protein by de novo sequencing using the combination of ProAlanase and tryptic peptides. Finally, we demonstrate that ProAlanase is efficient in disulfide bond mapping, showing high coverage of disulfide-containing regions in a non-reduced monoclonal antibody.
HostingRepository	PRIDE
AnnounceDate	2020-10-19
AnnouncementXML	Submission_2020-10-19_00:16:08.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Diana Samodova
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	acetylated residue; iodoacetamide derivatized residue; deamidated residue
Instrument	Q Exactive HF-X

Revision	Datetime	Status	ChangeLog Entry
0	2020-09-28 05:06:03	ID requested
⏵ 1	2020-10-19 00:16:09	announced

Samodova D, Hosfield CM, Cramer CN, Giuli MV, Cappellini E, Franciosa G, Rosenblatt MM, Kelstrup CD, Olsen JV, ProAlanase is an Effective Alternative to Trypsin for Proteomics Applications and Disulfide Bond Mapping. Mol Cell Proteomics, 19(12):2139-2157(2020) [pubmed]

submitter keyword: Proteomics, ProAlanase, sequence coverage, phosphorylation, proline-rich, low-pH, disulfide bonds, histones, de novo sequencing

Jesper Velgaard Olsen
contact affiliation	Professor, Novo Nordisk Foundation Center for Protein Research, Copenhagen, Denmark
contact email	jesper.olsen@cpr.ku.dk
lab head
Diana Samodova
contact affiliation	University of Copenhagen, Novo Nordisk Foundation Center for Protein Research
contact email	diana.samodova@cpr.ku.dk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD021703
     1. Label: PRIDE project
     2. Name: ProAlanase is an effective alternative to trypsin for proteomics applications and disulfide bond mapping_part3