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Covalent inhibition revealed by the crystal structure of the caspase-8/p35 complex

G Xu1, M Cirilli, Y Huang

  • 1Department of Biochemistry, Weill Medical College of Cornell University, New York, New York, 10021 USA

Nature
|March 22, 2001
PubMed

Insights

Baculovirus p35 protein prevents apoptosis by inhibiting caspases. Structural and biochemical studies reveal a novel mechanism where p35 forms a covalent bond, blocking caspase activity and providing broad-spectrum protease inhibition.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Structural Biology

Background:

  • Apoptosis is a regulated process vital for multicellular organism development and homeostasis.
  • Baculovirus p35 protein is a known inhibitor of apoptosis through broad-spectrum caspase inhibition.

Purpose of the Study:

  • To elucidate the molecular mechanism of p35-mediated caspase inhibition.
  • To determine the crystal structure of p35 in complex with human caspase-8.

Main Methods:

  • X-ray crystallography to obtain the 3.0 Å structure of p35-caspase-8 complex.
  • Biochemical assays and mutagenesis studies to validate structural findings.

Main Results:

  • The structure revealed a covalent thioester linkage between p35 and the caspase-8 active site.
  • Cleavage induces conformational changes in p35, repositioning its N-terminus to block the active site.
  • Mutations at the p35 N-terminus abrogated inhibitory activity, confirming its role.

Conclusions:

  • p35 employs a novel mechanism of caspase inhibition involving covalent linkage and conformational changes.
  • The N-terminus of p35 is critical for stabilizing the inhibition and preventing intermediate hydrolysis.
  • Conserved contacts outside the active site contribute to p35's broad-spectrum inhibitory activity.

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