A quantitative proteomics design for systematic identification of protease cleavage events.
We present here a novel proteomics design for systematic identification of protease cleavage events by quantitative N-terminal proteomics, circumventing the need for time-consuming manual validation. We bypass the singleton detection problem of protease-generated neo-N-terminal peptides by introducing differential isotopic proteome labeling such that these substrate reporter peptides are readily distinguished from all other N-terminal peptides. Our approach was validated using the canonical human caspase-3 protease and further applied to mouse cathepsin D and E substrate processing in a mouse dendritic cell proteome, identifying the largest set of protein protease substrates ever reported and gaining novel insight into substrate specificity differences of these cathepsins.
Impens
Francis
F
Department of Medical Protein Research, VIB, B-9000 Ghent, Belgium.
Colaert
Niklaas
N
Helsens
Kenny
K
Ghesquière
Bart
B
Timmerman
Evy
E
De Bock
Pieter-Jan
PJ
Chain
Benjamin M
BM
Vandekerckhove
Joël
J
Gevaert
Kris
K
D000595:Amino Acid Sequence; D000818:Animals; D053148:Caspase 3 / Q000378:metabolism*; D002402:Cathepsin D / Q000378:metabolism; D020059:Cathepsin E / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D051379:Mice; D008969:Molecular Sequence Data; D040901:Proteomics; D013379:Substrate Specificity
D053148:Caspase 3; D020059:Cathepsin E; D002402:Cathepsin D