computational prediction and experimental verification of new map kinase docking sites and substrates including gli transcription factors计算预测和实验验证新的map激酶对接网站和基质包括gli转录因子.pdfVIP

Computational Prediction and Experimental Verification of New MAP Kinase Docking Sites and Substrates Including Gli Transcription Factors 1,2,3 1,2 2 1,3 1 Thomas C. Whisenant , David T. Ho , Ryan W. Benz , Jeffrey S. Rogers , Robyn M. Kaake , 1,3 1 1,2,3 1,2,3 Elizabeth A. Gordon , Lan Huang , Pierre Baldi , Lee Bardwell * 1 Department of Developmental and Cell Biology, University of California, Irvine, California, United States of America, 2 Institute for Genomics and Bioinformatics, University of California, Irvine, California, United States of America, 3 Center for Complex Biological Systems, University of California, Irvine, California, United States of America Abstract In order to fully understand protein kinase networks, new methods are needed to identify regulators and substrates of kinases, especially for weakly expressed proteins. Here we have developed a hybrid computational search algorithm that combines machine learning and expert knowledge to identify kinase docking sites, and used this algorithm to search the human genome for novel MAP kinase substrates and regulators focused on the JNK family of MAP kinases. Predictions were tested by peptide array followed by rigorous biochemical verification with in vitro binding and kinase assays on wild-type and mutant proteins. Using this procedure, we found new ‘D-site’ class docking sites in previously known JNK substrates (hnRNP-K, PPM1J/PP2Czeta), as well as new JNK-interacting proteins (MLL4, NEIL1). Finally, we identified new D-site- dependent MAPK substrates, inc