An integrated high-performance liquid chromatography-mass spectrometry system for the activity-dependent analysis of matrix metalloproteases

Robert Freije, Theo Klein, Bert Ooms, Henk F. Kauffman, Rainer Bischoff*

*Bijbehorende auteur voor dit werk

OnderzoeksoutputAcademicpeer review

20 Citaten (Scopus)

Samenvatting

Matrix metalloproteases (MMPs) comprise a family of enzymes that play important roles in mediating angiogenesis, the remodelling of tissues and in cancer metastasis. Consequently, they are attractive targets for therapeutic intervention in chronic inflammation, cancer and neurological disorders. In order to study MMPs in body fluids in an activity-dependent manner, we have developed an automated, integrated system comprising an immobilized inhibitor cartridge for activity-dependent enrichment, an immobilized trypsin reactor for rapid on-line proteolysis and a capillary or nanoLC-MS system for separation and identification of the obtained peptide fragments. This targeted proteomics system was optimized with respect to recovery and evaluated through the analysis of urine samples that were spiked with recombinant MMP-12. MMP-12 specific peptide fragments were easily detected in a nanoLC-MS analysis of 500 mu L crude urine spiked at a level of 8 nM. These results show the feasibility of selective, activity-dependent enrichment of MMPs from a non-treated biofluid at low nM concentrations. (c) 2007 Elsevier B.V. All rights reserved.

Originele taal-2English
Pagina's (van-tot)417-425
Aantal pagina's9
TijdschriftJournal of Chromatography A
Volume1189
Nummer van het tijdschrift1-2
DOI's
StatusPublished - 2-mei-2008

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