Because of the multiplicity and broad substrate specificity of signaling enzymes, it is of immense importance to understand how the cell achieves efficiency and accuracy in signaling. post-synaptic density, disc large, zo-1 proteinĬells use a multitude of signaling proteins to alter cellular behavior in response to changes in their external and internal environment.non-catalytic region of tyrosine kinase adaptor protein.Fas-associated protein with death domain.dual-adapter for phosphotyrosine and 3-phosphoinositides.Thus, the field is in a state of continuous advance and expansion, which demands that the classification scheme be regularly updated and, if needed, revised. In addition, although not usually considered as scaffolds, several other proteins, such as regulatory proteins with catalytic activity, phosphatase targeting subunits, E3 ubiquitin ligases, ESCRT proteins in endosomal sorting and DNA damage sensors also function by bona fide scaffolding mechanisms. It will also be shown, however, that the categories partially overlap and often their names are used interchangeably in the literature. Here we discuss the categories of scaffolds, anchors, docking proteins and adaptors in some detail, and using many examples we demonstrate that they cover a wide range of functional modes that appear to segregate into three practical categories, simple proteins binding two partners together (adaptors), larger multidomain proteins targeting and regulating more proteins in complex ways (scaffold/anchoring proteins) and proteins specialized to initiate signaling cascades by localizing partners at the cell membrane (docking proteins). By binding and bringing into proximity two or more signaling proteins, these proteins direct the flow of information in the cell by activating, coordinating and regulating signaling events in regulatory networks. These high-affinity and isoform-specific inhibitors will enhance mechanistic and cellular investigations of SUMO biology.In this series of four minireviews the field of scaffold proteins and proteins of similar molecular/cellular functions is overviewed. The monobodies inhibited SUMO1/SIM interactions and, unexpectedly, also inhibited SUMO1 conjugation. By virtue of their ability to be designed to bind any protein of interest, OptoMBs have the potential to find new powerful applications as light-switchable binders of untagged proteins with the temporal and spatial precision afforded by = values of approximately 100 nM but bound to SUMO2 400 times more weakly. coli extract, achieving 99.8% purity and over 40% yield in a single purification step. We demonstrate that our αSH2-OptoMB can be used to purify SH2-tagged proteins directly from crude E. Here we present, as proof-of-principle, the development of a light-controlled monobody (OptoMB) that works in vitro and in cells and whose affinity for its SH2-domain target exhibits a 330-fold shift in binding affinity upon illumination. The ability to reversibly control their binding activity to their targets on demand would significantly expand their applications in biotechnology, medicine, and research. ![]() Abstract Monobodies are synthetic non-immunoglobulin customizable protein binders invaluable to basic and applied research, and of considerable potential as future therapeutics and diagnostic tools.
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