Key feature of the catalytic cycle of TNF-α converting enzyme involves communication between distal protein sites and the enzyme catalytic core

Ariel Solomon, Barak Akabayov, Anatoly Frenkel, Marcos E. Milla, Irit Sagi

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

Despite their key roles in many normal and pathological processes, the molecular details by which zinc-dependent proteases hydrolyze their physiological substrates remain elusive. Advanced theoretical analyses have suggested reaction models for which there is limited and controversial experimental evidence. Here we report the structure, chemistry and lifetime of transient metal-protein reaction intermediates evolving during the substrate turnover reaction of a metalloproteinase, the tumor necrosis factor-α converting enzyme (TACE). TACE controls multiple signal transduction pathways through the proteolytic release of the extracellular domain of a host of membrane-bound factors and receptors. Using stopped-flow x-ray spectroscopy methods together with transient kinetic analyses, we demonstrate that TACE's catalytic zinc ion undergoes dynamic charge transitions before substrate binding to the metal ion. This indicates previously undescribed communication pathways taking place between distal protein sites and the enzyme catalytic core. The observed charge transitions are synchronized with distinct phases in the reaction kinetics and changes in metal coordination chemistry mediated by the binding of the peptide substrate to the catalytic metal ion and product release. Here we report key local charge transitions critical for proteolysis as well as long sought evidence for the proposed reaction model of peptide hydrolysis. This study provides a general approach for gaining critical insights into the molecular basis of substrate recognition and turnover by zinc metalloproteinases that may be used for drug design.

Original languageEnglish
Pages (from-to)4931-4936
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume104
Issue number12
DOIs
StatePublished - 20 Mar 2007
Externally publishedYes

Keywords

  • Dynamics
  • Matrix metalloproteinases
  • Proteolysis
  • Stopped flow
  • X-ray absorption

Fingerprint

Dive into the research topics of 'Key feature of the catalytic cycle of TNF-α converting enzyme involves communication between distal protein sites and the enzyme catalytic core'. Together they form a unique fingerprint.

Cite this