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A quantitative proteomics design for systematic identification of protease cleavage events.

ABSTRACT: Recombinant mouse cathepsin D was added to lysates of mouse dendritic cells. Neo-N-terminal peptides were isolated by N-terminal COFRADIC and their presence in ratios distributed distinct from other peptides was induced by mixing light and heavy labeled proteins according to a novel quantitative design.

INSTRUMENT(S): LTQ Orbitrap, instrument model

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

SUBMITTER: Francis Impens

PROVIDER: PRD000281 | Pride | 2010-08-03

REPOSITORIES: Pride

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			Action	DRS
	PRIDE_Exp_Complete_Ac_13337.pride.mgf.gz	Mgf
	PRIDE_Exp_Complete_Ac_13337.pride.mztab.gz	Mztab
	PRIDE_Exp_Complete_Ac_13337.xml.gz	Xml
	PRIDE_Exp_Complete_Ac_13338.pride.mgf.gz	Mgf
	PRIDE_Exp_Complete_Ac_13338.pride.mztab.gz	Mztab

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Project description:Proteins' N-termini contain information about their biochemical properties and functions, and they can undergo co- or post-translational modifications, as well as be processed by proteases. To expand the coverage of the N-terminome, we have developed LATE (LysN Amino Terminal Enrichment), a method that uses selective chemical derivatization of α-amines to isolate the N-terminal peptides. We applied LATE in combination with other N-terminomic method to study caspase-3 mediated proteolysis both in vitro and during apoptosis in cells. This enable us to identify many unreported caspase-3 cleavages, including some that cannot be identified by other methods. Furthermore, we find direct evidence that some of the caspase-3 generated neo-N-termini can be further modified by Nt-acetylation. This provides a global picture of the caspase-3 degradome and uncovers previously unrecognised functional crosstalk between post-translational Nt-acetylation and caspase proteolysis pathways.

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