Substitution of methionine 435 with leucine, isoleucine, and serine in tumor necrosis factor alpha converting enzyme inactivates ectodomain shedding activity

Liliana Pérez, John E. Kerrigan, Xiaojin Li, Huizhou Fan

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

Tumor necrosis factor alpha (TNF-α) converting enzyme (TACE) is a zinc metalloprotease that has emerged as a general sheddase, which is responsible for ectodomain release of numerous membrane proteins, including the proinflammatory cytokine TNF-α, the leukocyte adhesin L-selectin and epidermal growth factor receptor ligand-transforming growth factor α (TGF-α), and related family members. Structurally, TACE belongs to a large clan of proteases, designated the metzincins, because TACE possesses a conserved methionine (Met435), frequently referred to as the met-turn residue, in its active site. A vital role of this residue in the function of TACE is supported by the fact that cells expressing the M435I TACE variant are defective in ectodomain shedding. However, the importance of Met435 in TACE appears to be uncertain, since another metzincin, matrix metalloprotease-2, has been found to be enzymatically fully active with either leucine or serine in place of its met-turn residue. We constructed TACE mutants with leucine or serine in place of Met435 to further examine the role of the met-turn residue in TACE-mediated ectodomain cleavage. Similar to the M435I TACE mutant, both the M435L and M435S constructs are defective in cleaving transmembrane TNF-α, TGF-α, and L-selectin. Comparative modeling and dynamic computation detected structural perturbations, which resulted in higher energy, in the catalytic zinc complexes of the Met435 TACE mutants compared with that in the wild-type enzyme. Thus, Met435 serves to maintain the stability of the catalytic center of TACE for the hydrolysis of peptide bonds in substrates.

Original languageEnglish (US)
Pages (from-to)141-149
Number of pages9
JournalBiochemistry and Cell Biology
Volume85
Issue number1
DOIs
StatePublished - Feb 2007

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Keywords

  • L-selectin
  • Met-turn
  • Metalloprotease
  • Metzincin
  • TACE
  • TGF-α
  • TNF-α

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