Kininogen-derived peptides for investigating the putative vasoactive properties of human cathepsins K and L.
Macrophages at an inflammatory site release massive amounts of proteolytic enzymes, including lysosomal cysteine proteases, which colocalize with their circulating, tight-binding inhibitors (cystatins, kininogens), so modifying the protease/antiprotease equilibrium in favor of enhanced proteolysis. We have explored the ability of human cathepsins B, K and L to participate in the production of kinins, using kininogens and synthetic peptides that mimic the insertion sites of bradykinin on human kininogens. Although both cathepsins processed high-molecular weight kininogen under stoichiometric conditions, only cathepsin L generated significant amounts of immunoreactive kinins. Cathepsin L exhibited higher specificity constants (kcat/Km) than tissue kallikrein (hK1), and similar Michaelis constants towards kininogen-derived synthetic substrates. A 20-mer peptide, whose sequence encompassed kininogen residues Ile376 to Ile393, released bradykinin (BK; 80%) and Lys-bradykinin (20%) when incubated with cathepsin L. By contrast, cathepsin K did not release any kinin, but a truncated kinin metabolite BK(5-9) [FSPFR(385-389)]. Accordingly cathepsin K rapidly produced BK(5-9) from bradykinin and Lys-bradykinin, and BK(5-8) from des-Arg9-bradykinin, by cleaving the Gly384-Phe385 bond. Data suggest that extracellular cysteine proteases may participate in the regulation of kinin levels at inflammatory sites, and clearly support that cathepsin K may act as a potent kininase.
Desmazes
Claire
C
Laboratoire d'Enzymologie et Chimie des Protéines, Equipe Protéases et Vectorisation, INSERM EMI-U 00 10, Université François Rabelais, Faculté de Médecine, Tours, France.
Galineau
Laurent
L
Gauthier
Francis
F
Brömme
Dieter
D
Lalmanach
Gilles
G
D000595:Amino Acid Sequence; D001920:Bradykinin / Q000737:chemistry / Q000378:metabolism; D002401:Cathepsin B / Q000378:metabolism; D056657:Cathepsin K; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism; D006801:Humans; D007704:Kininogens / Q000378:metabolism*; D007705:Kinins / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism*; D020840:Tissue Kallikreins / Q000737:chemistry / Q000378:metabolism; D014664:Vasodilation / Q000502:physiology
D007704:Kininogens; D007705:Kinins; D010446:Peptide Fragments; D002403:Cathepsins; D020840:Tissue Kallikreins; D003546:Cysteine Endopeptidases; D002401:Cathepsin B; C534281:CTSL protein, human; D056668:Cathepsin L; C534130:CTSK protein, human; D056657:Cathepsin K; D001920:Bradykinin