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Profiling constitutive proteolytic events in vivo.

The Biochemical journal (2007-07-26)
John C Timmer, Mari Enoksson, Eric Wildfang, Wenhong Zhu, Yoshinobu Igarashi, Jean-Benard Denault, Yuliang Ma, Benjamin Dummitt, Yie-Hwa Chang, Alan E Mast, Alexey Eroshkin, Jeffrey W Smith, W Andy Tao, Guy S Salvesen
RESUMO

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 aminopeptidase 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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O-Methylisourea hemisulfate salt, 99%
Sigma-Aldrich
o-Methylisourea bisulfate, 99%