Tài liệu Báo cáo khoa học: Plasticity of S2–S4 specificity pockets of executioner caspase-7 revealed

Plasticity of S2–S4 specificity pockets of executioner

caspase-7 revealed by structural and kinetic analysis

Johnson Agniswamy, Bin Fang and Irene T. Weber

Department of Biology, Molecular Basis of Disease, Georgia State University, Atlanta, GA, USA

Caspases, the key effector molecules in apoptosis, are

potential targets for pharmacological modulation of

cell death. Uncontrolled apoptosis due to enhanced

caspase activity occurs in nerve crush injury, stroke

and neurodegenerative diseases such as Alzheimer’s,

Parkinson’s and Huntington’s diseases [1–3]. On the

other hand, inadequate caspase activity is implicated in

cancer, autoimmune diseases and viral infections [4–6],

and a number of potential drugs are being developed

for selective induction of apoptosis in cancer cells [7,8].

The substrate-based peptide inhibitor zVAD-fmk pro￾vides substantial protection against stroke, myocardial

infarction, osteoarthritis, hepatic injury, sepsis, and

amyotrophic lateral sclerosis in animal models [9–11].

Small nonpeptide inhibitors are preferred for their

superior metabolic stability and cell permeability. Two

nonpeptide inhibitors are currently in phase II clinical

trials: IDN6556 for treatment of acute-tissue injury

disease and liver diseases [12], and VX-740 for treat￾ment of rheumatoid arthritis [13]. Knowledge of the

molecular basis for substrate specificity of caspases is

critical for design of therapeutic agents for selective

control of cell death.

Caspases are cysteine proteases that hydrolyze the

peptide bond after an aspartate residue [14–17]. Thir￾teen human caspases have been cloned and character￾ized to varying extents [18,19]. Caspases are classified

into three groups based on their function and

Keywords

allosteric site; apoptosis; cysteine protease;

enzyme

Correspondence

I. T. Weber, Department of Biology, Georgia

State University, PO Box 4010, Atlanta,

GA 30302, USA

Fax: +1 404 413 5301

Tel: +1 404 413 5411

E-mail: [email protected]

Database

The atomic coordinates and structure

factors have been deposited in the Protein

Data Bank under the accession codes 2QL5

for caspase-7 ⁄ DMQD, 2QL9 for caspase-7 ⁄

DQMD, 2QLF for caspase-7 ⁄ DNLD, 2QL7

for caspase-7 ⁄ IEPD, 2QLB for caspase-7 ⁄

ESMD, and 2QLJ for caspase-7 ⁄ WEHD

(Received 18 April 2007, revised 3 July

2007, accepted 17 July 2007)

doi:10.1111/j.1742-4658.2007.05994.x

Many protein substrates of caspases are cleaved at noncanonical sites in

comparison to the recognition motifs reported for the three caspase sub￾groups. To provide insight into the specificity and aid in the design of

drugs to control cell death, crystal structures of caspase-7 were determined

in complexes with six peptide analogs (Ac-DMQD-Cho, Ac-DQMD-Cho,

Ac-DNLD-Cho, Ac-IEPD-Cho, Ac-ESMD-Cho, Ac-WEHD-Cho) that

span the major recognition motifs of the three subgroups. The crystal

structures show that the S2 pocket of caspase-7 can accommodate diverse

residues. Glu is not required at the P3 position because Ac-DMQD-Cho,

Ac-DQMD-Cho and Ac-DNLD-Cho with varied P3 residues are almost as

potent as the canonical Ac-DEVD-Cho. P4 Asp was present in the better

inhibitors of caspase-7. However, the S4 pocket of executioner caspase-7

has alternate regions for binding of small branched aliphatic or polar resi￾dues similar to those of initiator caspase-8. The observed plasticity of the

caspase subsites agrees very well with the reported cleavage of many pro￾teins at noncanonical sites. The results imply that factors other than the

P4–P1 sequence, such as exosites, contribute to the in vivo substrate speci￾ficity of caspases. The novel peptide binding site identified on the molecular

surface of the current structures is suggested to be an exosite of caspase-7.

These results should be considered in the design of selective small molecule

inhibitors of this pharmacologically important protease.

Abbreviation

PARP, poly(ADP-ribose) polymerase.

4752 FEBS Journal 274 (2007) 4752–4765 ª 2007 The Authors Journal compilation ª 2007 FEBS