Web and database software for identification of intact proteins using "top down" mass spectrometry.
For the identification and characterization of proteins harboring posttranslational modifications (PTMs), a "top down" strategy using mass spectrometry has been forwarded recently but languishes without tailored software widely available. We describe a Web-based software and database suite called ProSight PTM constructed for large-scale proteome projects involving direct fragmentation of intact protein ions. Four main components of ProSight PTM are a database retrieval algorithm (Retriever), MySQL protein databases, a file/data manager, and a project tracker. Retriever performs probability-based identifications from absolute fragment ion masses, automatically compiled sequence tags, or a combination of the two, with graphical rendering and browsing of the results. The database structure allows known and putative protein forms to be searched, with prior or predicted PTM knowledge used during each search. Initial functionality is illustrated with a 36-kDa yeast protein identified from a processed cell extract after automated data acquisition using a quadrupole-FT hybrid mass spectrometer. A +142-Da delta(m) on glyceraldehyde-3-phosphate dehydrogenase was automatically localized between Asp90 and Asp192, consistent with its two cystine residues (149 and 153) alkylated by acrylamide (+71 Da each) during the gel-based sample preparation. ProSight PTM is the first search engine and Web environment for identification of intact proteins (https://prosightptm.scs.uiuc.edu/).
Taylor
Gregory K
GK
Department of Computer Science, University of Illinois, Urbana, Illinois 61820, USA.
Kim
Yong-Bin
YB
Forbes
Andrew J
AJ
Meng
Fanyu
F
McCarthy
Ryan
R
Kelleher
Neil L
NL
D000595:Amino Acid Sequence; D000818:Animals; D030562:Databases, Protein; D020407:Internet; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D020539:Sequence Analysis, Protein; D016324:Sequence Tagged Sites; D012984:Software
D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins