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Title 

SPR imaging-based monitoring of caspase-3 activation

Authors 

Kyoungsook ParkJun Hyoung AhnSo Yeon YiMoonil KimBong Hyun Chung

Publisher 

Elsevier

Issue Date 

2008

Citation 

Biochemical and Biophysical Research Communications, vol. 368, no. 3, pp. 684-689

Keywords 

caspase-3 activationlinker peptideprotein chipSPR imaging systemcaspase 3enzyme activationimaging system

Abstract 

The activation of caspase-3 plays an important role in the apoptotic process. In this study, we describe a novel method by which caspase-3-dependent proteolytic cleavage can be monitored, using a surface plasmon resonance (SPR) imaging protein chip system. To the best of our knowledge, this is the first report regarding the SPR imaging-based monitoring of caspase-3 activation. In order to evaluate the performance of this protocol, we constructed a chimeric caspase-3 substrate (GST:DEVD:EGFP) comprised of glutathione S transferase (GST) and enhanced green fluorescent protein (EGFP) with a specialized linker peptide harboring the caspase-3 cleavage sequence, DEVD. Using this reporter, we assessed the cleavage of the artificial caspase-3 substrate in response to caspase-3 using an SPR imaging sensor. The purified GST:DEVD:EGFP protein was initially immobilized onto a glutathionylated gold chip surface, and subsequently analyzed using an SPR imaging system. As a result, caspase-3 activation predicated on the proteolytic properties inherent to substrate specificity could be monitored via an SPR imaging system with a detection performance similar to that achievable by the conventional method, including fluorometric assays. Collectively, our data showed that SPR imaging protein chip system can be effectively utilized to monitor the proteolytic cleavage in caspase-3, thereby potentially enabling the detection of other intracellular protease activation via the alteration of the protease recognition site in the linker peptides.

ISSN 

0006-291X

Link 

http://dx.doi.org/10.1016/j.bbrc.2008.01.137

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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