Journal article

An Optimized Activity-Based Probe for the Study of Caspase-6 Activation

Laura E Edgington, Bram J van Raam, Martijn Verdoes, Christoph Wierschem, Guy S Salvesen, Matthew Bogyo

CHEMISTRY & BIOLOGY | CELL PRESS | Published : 2012

Abstract

Although significant efforts have been made to understand the mechanisms of caspase activation during apoptosis, many questions remain regarding how and when executioner caspases get activated. We describe the design and synthesis of an activity-based probe that labels caspase-3/-6/-7, allowing direct monitoring of all executioner caspases simultaneously. This probe has enhanced in vivo properties and reduced cross-reactivity compared to our previously reported probe, AB50. Using this probe, we find that caspase-6 undergoes a conformational change and can bind substrates even in the absence of cleavage of the proenzyme. We also demonstrate that caspase-6 activation does not require active ca..

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Awarded by NIH


Awarded by NATIONAL CANCER INSTITUTE


Awarded by NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES


Awarded by NATIONAL INSTITUTE OF BIOMEDICAL IMAGING AND BIOENGINEERING


Funding Acknowledgements

We thank D. Ehrnhoefer, N. Skotte, and M. Hayden of the University of British Columbia for critical discussions of caspase-6 biology, C. Pop, formerly of the Sanford-Burnham Medical Research Institute, for helpful advice on the enzyme kinetics experiments, and S. Snipas for his help in cloning the caspase-6 mutants. We also thank T. Doyle at the Stanford Small Animal Facility for assistance with the optical imaging studies. This work was supported by a grant from the NIH (R01 EB005011). B.J.v.R. and M.V. are supported by a Rubicon fellowship from the Netherlands Organization for Scientific Research, and B.J.v.R. by an additional fellowship from the Barth Syndrome Foundation.