Identification of mouse liver proteins on two‐dimensional electrophoresis gels by matrix‐assisted laser desorption/ionization mass spectrometry ofin situenzymatic digests
作者:
Kathy L. O'Connell,
John T. Stults,
期刊:
ELECTROPHORESIS
(WILEY Available online 1997)
卷期:
Volume 18,
issue 3‐4
页码: 349-359
ISSN:0173-0835
年代: 1997
DOI:10.1002/elps.1150180309
出版商: Wiley Subscription Services, Inc., A Wiley Company
关键词: Two‐dimensional polyacrylamide gel electrophoresis;Matrix‐assisted laser desorption;ionization – time of flight;Peptide mass mapping;Protein identification
数据来源: WILEY
摘要:
AbstractA number of proteins from a silver‐stained two‐dimensional (2‐D) electrophoresis gel of mouse liver whole‐cell lysate were identified by peptide mass mapping and sequene database searching. The excised protein spots were processed byin situreduction and alkylation, followed by Lys‐C digestion. The masses of the resulting peptide mixtures were measured with a matrix‐assisted laser desorption/ionization (MALDI) reflectron‐time‐of‐flight mass spectrometer. These masses were used successfully to search a protein sequence database. Optimized silver staining and digestion protocols allowed proteins to be identified routinely at the low picomole level. The high mass accuracy and resolution provided by delayed extraction were important for high specificity in the database search. Fragment ion data obtained by MALDI post‐source decay (PSD) measurements not only provided confirmation of peptide identification, but could be used to identify the protein from a single peptide without spec
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