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Structural basis for executioner caspase recognition of P5 position in substrates

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Abstract

Caspase-3, -6 and -7 cleave many proteins at specific sites to induce apoptosis. Their recognition of the P5 position in substrates has been investigated by kinetics, modeling and crystallography. Caspase-3 and -6 recognize P5 in pentapeptides as shown by enzyme activity data and interactions observed in the crystal structure of caspase-3/LDESD and in a model for caspase-6. In caspase-3 the P5 main-chain was anchored by interactions with Ser209 in loop-3 and the P5 Leu side-chain interacted with Phe250 and Phe252 in loop-4 consistent with 50% increased hydrolysis of LDEVD relative to DEVD. Caspase-6 formed similar interactions and showed a preference for polar P5 in QDEVD likely due to interactions with polar Lys265 and hydrophobic Phe263 in loop-4. Caspase-7 exhibited no preference for P5 residue in agreement with the absence of P5 interactions in the caspase-7/LDESD crystal structure. Initiator caspase-8, with Pro in the P5-anchoring position and no loop-4, had only 20% activity on tested pentapeptides relative to DEVD. Therefore, caspases-3 and -6 bind P5 using critical loop-3 anchoring Ser/Thr and loop-4 side-chain interactions, while caspase-7 and -8 lack P5-binding residues.

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Abbreviations

Ac:

Acetyl

CHO:

Aldehyde

pNA:

p-Nitroanilide

RMS:

Root mean square

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Acknowledgements

G.F. and B·F. were supported in part by the Georgia State University Research Program Enhancement award. B·F. was supported by the Georgia State University Molecular Basis of Disease Fellowship. I.T.W. and R.W·H. are Georgia Cancer Coalition Distinguished Cancer Scholars. This research was supported in part by the Georgia Research Alliance, the Georgia Cancer Coalition and the National Institutes of Health award GM065762. We thank the staff at the SER-CAT beamline at the Advanced Photon Source, Argonne National Laboratory, for assistance during X-ray data collection. Use of the Advanced Photon Source was supported by the U. S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. DE-AC02-06CH11357.

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Authors and Affiliations

  1. Department of Biology, Molecular Basis of Disease Program, Georgia State University, Atlanta, GA, 30303, USA

    Guoxing Fu, Alexander A. Chumanevich, Johnson Agniswamy, Bin Fang & Irene T. Weber

  2. Department of Biology, Georgia State University, P.O. Box 4010, Atlanta, GA, 30302, USA

    Guoxing Fu, Alexander A. Chumanevich, Johnson Agniswamy, Bin Fang & Irene T. Weber

  3. Department of Computer Science, Molecular Basis of Disease Program, Georgia State University, Atlanta, GA, 30303, USA

    Robert W. Harrison

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  1. Guoxing Fu
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Correspondence to Irene T. Weber.

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Fu, G., Chumanevich, A.A., Agniswamy, J. et al. Structural basis for executioner caspase recognition of P5 position in substrates. Apoptosis 13, 1291–1302 (2008). https://doi.org/10.1007/s10495-008-0259-9

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