PDBsum entry: 1i4e

Apoptosis/hydrolase

PDB id: 1i4e

Contents

Protein chains

294 a.a. *

243 a.a. *

* Residue conservation analysis

PDB id:

1i4e

Name:

Apoptosis/hydrolase

Title:

Crystal structure of the caspase-8/p35 complex

Structure:

Early 35 kda protein. Chain: a. Synonym: p35, apoptosis-preventing protein. Engineered: yes. Caspase-8. Chain: b. Synonym: casp-8, ice-like apoptotic protease 5, mort1-assoc 3 homolog, mach, fadd-homologous ice/ced-3-like protease, f ice, flice, apoptotic cysteine protease, apoptotic protease

Source:

Autographa californica nucleopolyhedro acmnpv. Organism_taxid: 46015. Gene: p35. Expressed in: escherichia coli. Expression_system_taxid: 562. Homo sapiens. Human. Organism_taxid: 9606.

Biol. unit:

Tetramer (from PQS)

Resolution:

3.00Å R-factor: 0.236 R-free: 0.296

Authors:

G.Xu,M.Cirilli,Y.Huang,R.L.Rich,D.G.Myszka,H.Wu

Key ref:

G.Xu et al. (2001). Covalent inhibition revealed by the crystal structure of the caspase-8/p35 complex. Nature, 410, 494-497. PubMed id: 11260720 DOI: 10.1038/35068604

Date:

20-Feb-01 Release date: 28-Mar-01

Protein chain

P08160  (VP35_NPVAC) -  Early 35 kDa protein

Seq: 299 a.a.

Struc: 293 a.a.

Protein chain

Q14790  (CASP8_HUMAN) -  Caspase-8

Seq: 479 a.a.

Struc: 243 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: Chain B: E.C.3.4.22.61 - Caspase-8.

 Gene Ontology (GO) functional annotation 

• Biological process: apoptosis (3 terms)
• Biochemical function: protein binding (6 terms)

DOI no: 10.1038/35068604

Nature 410:494-497 (2001)

PubMed id: 11260720

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

G.Xu, M.Cirilli, Y.Huang, R.L.Rich, D.G.Myszka, H.Wu.

ABSTRACT

Apoptosis is a highly regulated process that is crucial for normal development and homeostasis of multicellular organisms. The p35 protein from baculoviruses effectively prevents apoptosis by its broad-spectrum caspase inhibition. Here we report the crystal structure of p35 in complex with human caspase-8 at 3.0 A resolution, and biochemical and mutagenesis studies based on the structural information. The structure reveals that the caspase is inhibited in the active site through a covalent thioester linkage to p35, which we confirmed by gel electrophoresis, hydroxylamine treatment and mass spectrometry experiments. The p35 protein undergoes dramatic conformational changes on cleavage by the caspase. The repositioning of the amino terminus of p35 into the active site of the caspase eliminates solvent accessibility of the catalytic dyad. This may be crucial for preventing hydrolysis of the thioester intermediate, which is supported by the abrogation of inhibitory activity through mutations at the N terminus of p35. The p35 protein also makes conserved contacts with the caspase outside the active-site region, providing the molecular basis for the broad-spectrum inhibitory activity of this protein. We demonstrate a new molecular mechanism of caspase inhibition, as well as protease inhibition in general.

Selected figure(s)

Figure 1.
Figure 1: Structure of the p35/caspase-8 complex. a, Ribbon diagram of dimeric complex with the two-fold axis in the vertical orientation. p35, cyan and green; -subunit (p18) of caspase-8, magenta and red; -subunit (p12) of caspase-8, orange and yellow. Ordered termini for p35-N (residues 2 -87) and p35-C (residues 93 -299) are labelled. b, Conformational transitions of p35 on cleavage. Residues with differences in C positions larger than 4.0 Å are shown in red, which include the N terminus (residues 2 -12), the CD loop (residues 35 -40), the caspase recognition sequence (residues 85 -87), the reactive-site loop after the cleavage site (residues 93 -101), the FG loop (residues 157 -165) and the KL loop (residues 254 -255). c, Atomic model of the complex near the active site of caspase-8 overlaid with an omit electron density map (1.0 contour). Potential hydrogen bonds are indicated by dotted lines. Side chains for residue Met 86 of p35 and Tyr 412 of caspase-8 are omitted for clarity.

Figure 3.
Figure 3: Detailed interaction between p35 and caspase-8. a, Interaction near the N terminus of p35 with the active site of caspase-8. p35, cyan; -subunit of caspase-8, magenta. b, Footprint of p35 on the surface of caspase-8 (grey and yellow for each of the -units). Residues conserved among different caspases and main-chain atoms are shown in blue, whereas those with significant variations are shown in red. c, Interaction near the KL loop of p35. p35, cyan; -subunit of caspase-8, orange.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nature (2001, 410, 494-497) copyright 2001.

Figures were selected by the author.