Selective mitogen activated protein kinase activity sensors through the application of directionally programmable D domain motifs.

TitleSelective mitogen activated protein kinase activity sensors through the application of directionally programmable D domain motifs.
Publication TypeJournal Article
Year of Publication2014
AuthorsPeterson, LB, Yaffe, MB, Imperiali, B
Date Published2014 Sep 16
KeywordsAmino Acid Sequence, Biosensing Techniques, Blotting, Western, Mitogen-Activated Protein Kinases, Models, Molecular, Molecular Sequence Data, Sequence Homology, Amino Acid

Accurate and quantitative methods for measuring the dynamic fluctuations of protein kinase activities are critically needed as diagnostic tools and for the evaluation of kinase-targeted inhibitors, which represent a major therapeutic development area in the treatment of cancer and other diseases. In particular, rapid and economical methods that utilize simple instrumentation and provide quantitative data in a high throughput format will have the most impact on basic research in systems biology and medicine. There are over 500 protein kinases in the human kinome. Among these, the mitogen activated protein (MAP) kinases are recognized to be central players in key cellular signaling events and are associated with essential processes including growth, proliferation, differentiation, migration, and apoptosis. The major challenge with MAP kinase sensor development is achieving high selectivity since these kinases rely acutely on secondary interactions distal to the phosphorylation site to impart substrate specificity. Herein we describe the development and application of selective sensors for three MAP kinase subfamilies, ERK1/2, p38α/β, and JNK1/2/3. The new sensors are based on a modular design, which includes a sensing element that exploits a sulfonamido-oxine (Sox) fluorophore for reporting phosphorylation, a recognition and specificity element based on reported docking domain motifs and a variable linker, which can be engineered to optimize the intermodule distance and relative orientation. Following rigorous validation, the capabilities of the new sensors are exemplified through the quantitative analysis of the target MAP kinases in breast cancer progression in a cell culture model, which reveals a strong correlation between p38α/β activity and increased tumorgenicity.

Alternate JournalBiochemistry
PubMed ID25153342
PubMed Central IDPMC4165445
Grant ListES015339 / ES / NIEHS NIH HHS / United States
GM104047 / GM / NIGMS NIH HHS / United States
U54-CA112967 / CA / NCI NIH HHS / United States