Proteome-wide Identification of HtrA2/Omi Substrates.

Abstract

To identify apoptotic targets of HtrA2/Omi, we purified recombinant HtrA2/Omi and its catalytically inactive S306A mutant. Lysates of human Jurkat T lymphocytes incubated with either wild-type recombinant HtrA2/Omi or the S306A mutant were screened using the gel-free COFRADIC approach that isolates peptides covering the N-terminal parts of proteins. Analysis of the 1162 proteins identified by mass spectrometry yielded 15 HtrA2/Omi substrates of potential physiological relevance together holding a total of 50 cleavage sites. Several processing events were validated by incubating purified recombinant HtrA2/Omi with in vitro translated substrates or with Jurkat cell lysates. In addition, the generated set of cleavage sites was used to assess the protein substrate specificity of HtrA2/Omi. Our results suggest that HtrA2/Omi has a rather narrow cleavage site preference and that cytoskeletal proteins are prime targets of this protease.

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@article{Walle2007ProteomewideIO, title={Proteome-wide Identification of HtrA2/Omi Substrates.}, author={Lieselotte Vande Walle and Petra Van Damme and Mohamed Lamkanfi and Xavier Saelens and Jo{\"{e}l Vandekerckhove and Kris Gevaert and Peter Vandenabeele}, journal={Journal of proteome research}, year={2007}, volume={6 3}, pages={1006-15} }