Partial purification and characterization of a tumor necrosis factor-alpha converting activity.
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.
Robache-Gallea
S
S
Domaine Thérapeutique Immunologie, Roussel Uclaf, Romainville, France.
Bruneau
J M
JM
Robbe
H
H
Morand
V
V
Capdevila
C
C
Bhatnagar
N
N
Chouaib
S
S
Roman-Roman
S
S
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*
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