Journal
BIOCHEMICAL JOURNAL
Volume 407, Issue -, Pages 41-48Publisher
PORTLAND PRESS LTD
DOI: 10.1042/BJ20070775
Keywords
mass spectrometry; methionine aminopeptidase (MetAP); mitochondrial peptidase; N-terminal labelling; peptidase; protease; signal peptidase
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Funding
- NCRR NIH HHS [R21 RR019752, RR19752, U54 RR020843, RR20843] Funding Source: Medline
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Most known organisms encode proteases that are crucial for constitutive proteolytic events. In the present paper, we describe a method to define these events in proteomes from Escherichia coli to humans. The method takes advantage of specific N-terminal biotinylation of protein samples, followed by affinity enrichment and conventional LC (liquid chromatography)-MS/ MS (tandem mass spectrometry) analysis. The method is simple, uses conventional and easily obtainable reagents, and is applicable to most proteomics facilities. As proof of principle, we demonstrate profiles of proteolytic events that reveal exquisite in vivo specificity of methionine arninopeptidase in E. coli and unexpected processing of mitochondrial transit peptides in yeast, mouse and human samples. Taken together, our results demonstrate how to rapidly distinguish real proteolysis that occurs in vivo from the predictions based on in vitro experiments.
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