Profiling Activity

Science's STKE  27 Aug 2002:
Vol. 2002, Issue 147, pp. tw321-TW321
DOI: 10.1126/stke.2002.147.tw321

As scientists work to integrate the complex and vast information about cellular responses to the environment, new approaches are being developed to screen large, complex samples for changes that occur in response to cellular signal transduction cascades. Winssinger et al. describe a method for profiling multiple enzymatic activities from a cell lysate simultaneously. The method relies on creating a library of small molecule inhibitors and then being able to capture the ones that have interacted with the activated enzymes in the preparation and to visualize the abundance of the labeled small molecule inhibitor captured. They demonstrate a proof-of-principle experiment in which they can selectively detect the activation of caspase-3 during the initiation of apoptosis in Jurkat cells in response to granzyme B treatment. This methodology has several potential applications. First, small molecule inhibitors of known specificity can be used to determine the activity profile of defined targets. Second, in a cellular lysate, novel targets can be identified by changing the chemistry (linking the inhibitor to biotin, for example) such that the small molecule inhibitor and the activated enzyme can be copurified and the enzyme can then be subjected to sequence analysis. Finally, novel inhibitors can be discovered and then the small molecule inhibitor can be applied to cells to confirm its pharmacological activity.

N. Winssinger, S. Ficarro, P. G. Schultz, J. L. Harris, Profiling protein function with small molecule microarrays. Proc. Natl. Acad. Sci. U.S.A. 99, 11139-11144 (2002). [Abstract] [Full Text]