TY - JOUR T1 - An open library of human kinase domain constructs for automated bacterial expression JF - bioRxiv DO - 10.1101/038711 SP - 038711 AU - Daniel L. Parton AU - Sonya M. Hanson AU - Lucelenie Rodríguez-Laureano AU - Steven K. Albanese AU - Scott Gradia AU - Chris Jeans AU - Markus Seeliger AU - Nicholas Levinson AU - John D. Chodera Y1 - 2016/01/01 UR - http://biorxiv.org/content/early/2016/02/25/038711.abstract N2 - Kinases play a critical role in cellular signaling pathways. Human kinase dysregulation has been linked to a number of diseases, such as cancer, diabetes, and inflammation, and as a result, much of the effort in developing treatments (and perhaps 30%of all current drug development effort) has focused on shutting down aberrant kinases with targeted inhibitors. While insect and mammalian expression systems are frequently utilized for the expression of human kinases, they cannot compete with the simplicity and cost-effectiveness of bacterial expression systems, which historically had found human kinases difficult to express. Following the demonstration that phosphatase coexpression could give high yields of Src and Abl kinase domains in inexpensive bacterial expression systems [1], we have performed a large-scale expression screen to generate a library of His-tagged human kinase domain constructs that express well in a simple automated bacterial expression system where phosphatase coexpression (YopH for Tyr kinases, lambda for Ser/Thr kinases) is used. Starting from 96 kinases with crystal structures and any reported bacterial expression, we engineered a library of human kinase domain constructs and screened their coexpression with phosphatase, finding 52 kinases with yields greater than 2 mg/L culture. All sequences and expression data are provided online at https://github.com/choderalab/kinase-ecoli-expression-panel, and the plasmids are in the process of being made available through AddGene. ER -