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Mapping Domain Interaction Networks in Protein Kinases with Optical Tweezers
Author(s) -
Maillard Rodrigo
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.662.2
Subject(s) - optical tweezers , protein subunit , allosteric regulation , biophysics , chemistry , molecular switch , microbiology and biotechnology , biology , physics , biochemistry , molecule , receptor , gene , organic chemistry , quantum mechanics
Signaling proteins are dynamic molecular switches that sample multiple conformational states. Such conformational plasticity allows these proteins to adapt and respond to different biological signals. Protein Kinase A (PKA) is a signaling protein complex composed of regulatory and catalytic subunits. The regulatory subunit harbors two cAMP binding domains (A and B) that oscillate between inactive and active conformations depending on cAMP concentration. In this study, we use optical tweezers to identify the networks of communication between cAMP binding domains that enable PKA to switch efficiently and cooperatively from inactive to active states. We show that in the inactive PKA conformation, domain B of the regulatory subunit serves as an energetic hub, controlling the magnitude of interaction between domain A and the catalytic subunit, and whose mechanical perturbation triggers the highly coordinated dissociation of the PKA complex. In the presence of cAMP, domains A and B establish an intricate network of communication that involves direct, interfacial domain contacts as well as long‐range interactions between non‐contiguous structural motifs. In contrast, without ligand the two domains behave as independent, non‐interacting structures. Lastly, we demonstrate the application of optical tweezers to dissect how loss‐of‐function mutations can disrupt very distinct communication networks in PKA. The experimental approach based on optical tweezers should be readily applicable to study disease mutations, the mechanism of action of small molecules, or other signaling proteins. Support or Funding Information NSF MCB 1715572 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .