title: Partial purification and characterization of a tumor necrosis factor-alpha converting activity.
abstract: Tumor necrosis factor (TNF)-alpha is initially synthesized as an extracellular membrane-associated 26-kDa protein that is further cleaved at Ala76-Val77 to yield the soluble 17-kDa form. Recently, peptide-hydroxamate metalloproteinase inhibitors have been reported to block the proteolytic processing of TNF-alpha, thus suggesting that the putative TNF-alpha converting enzyme (TACE) is a zinc-dependent metalloendopeptidase. In this report, we characterize a TNF-alpha converting activity (TACA) that cleaves in vitro the human 26-kDa TNF-alpha at the physiological processing site. The chromatography steps followed for purification and the use of a panel of proteinase inhibitors indicate that the enzyme responsible for TACA is a membrane glycosylated metalloendopeptidase which is most likely different from the matrix-degrading metalloproteinases. The failure of TACA to process a Val77-->Gly77 precursor mutant emphasizes the importance of hydrophobic residue at P1' position. In addition, TACA is not able to cleave the mouse pro-TNF-alpha and does not catalyze in vitro the processing of other transmembrane proteins susceptible to metalloproteinase-mediated shedding, such as interleukin-6 or TNF receptors. These studies suggest the existence of an enzyme specific for TNF-alpha within the metalloproteinases involved in the processing/shedding of a number of cytokines and cytokine receptors.
authors.affiliation: Domaine Thérapeutique Immunologie, Roussel Uclaf, Romainville, France.
authors.forename: S
authors.initials: S
authors.lastname: Robache-Gallea
authors.forename: J M
authors.initials: JM
authors.lastname: Bruneau
authors.forename: H
authors.initials: H
authors.lastname: Robbe
authors.forename: V
authors.initials: V
authors.lastname: Morand
authors.forename: C
authors.initials: C
authors.lastname: Capdevila
authors.forename: N
authors.initials: N
authors.lastname: Bhatnagar
authors.forename: S
authors.initials: S
authors.lastname: Chouaib
authors.forename: S
authors.initials: S
authors.lastname: Roman-Roman
mesh_terms: D051722:ADAM Proteins; D000072198:ADAM17 Protein; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000302:isolation & purification; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D051379:Mice; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D006877:Hydroxamic Acids; D008562:Membrane Glycoproteins; C106339:RU 36156; D014409:Tumor Necrosis Factor-alpha; D051722:ADAM Proteins; D008666:Metalloendopeptidases; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human; C000606463:Adam17 protein, mouse