title: Covalent binding of 4-hydroxynonenal to matrix metalloproteinase 13 studied by liquid chromatography-mass spectrometry.
abstract: Osteoarthritis (OA) is caused by the degradation of articular cartilage and affects approximately 80% of people over the age of 65. Matrix metalloproteinases (MMPs) belong to a group of zinc endopeptidases that degrade extracellular matrix (ECM) proteins in cartilage. MMP-13, also known as collagenase 3, cleaves type II collagen more rapidly than other MMPs and therefore is an important target for the treatment of OA. The lipid peroxidation product 4-hydroxy-2-(E)-nonenal (HNE), generated under oxidative stress, is known to play a crucial role in cartilage degradation; however, the mechanism is not yet fully understood. An approach has been developed to monitor HNE modification sites by incubating rhMMP-13 ± HNE in vitro followed by analysis of tryptic digests by UHPLC coupled to high resolution (HR) quadrupole-time-of-flight (QqTOF) tandem mass spectrometry (MS/MS). The analysis elucidated several covalently modified histidine and cysteine residues. The reaction was monitored using different HNE concentrations and incubation times. A targeted assay, using multiple-reaction monitoring (MRM), was then optimized to increase the sensitivity of detecting these modification sites in biological samples. HNE-related covalent modifications of MMP-13 were confirmed in enriched extracts from interleukin 1β-activated chondrocytes from OA patients using HR-MS/MS and MRM analysis.
authors.affiliation: Chemistry Department/Pharmaqam, Université du Québec à Montréal , Montreal, Quebec H3C 3P8, Canada.
authors.forename: Makan
authors.initials: M
authors.lastname: Golizeh
authors.forename: Jamilah
authors.initials: J
authors.lastname: Abusarah
authors.forename: Mohamed
authors.initials: M
authors.lastname: Benderdour
authors.forename: Lekha
authors.initials: L
authors.lastname: Sleno
mesh_terms: D000447:Aldehydes / Q000737:chemistry*; D000595:Amino Acid Sequence; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D047468:Immunoprecipitation; D053583:Interleukin-1beta / Q000494:pharmacology; D053509:Matrix Metalloproteinase 13 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010003:Osteoarthritis / Q000378:metabolism / Q000473:pathology; D010455:Peptides / Q000032:analysis / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D053719:Tandem Mass Spectrometry
chemical_list: D000447:Aldehydes; D053583:Interleukin-1beta; D010455:Peptides; D011994:Recombinant Proteins; D053509:Matrix Metalloproteinase 13; C027576:4-hydroxy-2-nonenal