title: Designed phosphoprotein recognition in Escherichia coli.
abstract: Protein phosphorylation is a central biological mechanism for cellular adaptation to environmental changes. Dysregulation of phosphorylation signaling is implicated in a wide variety of diseases. Thus, the ability to detect and quantify protein phosphorylation is highly desirable for both diagnostic and research applications. Here we present a general strategy for detecting phosphopeptide-protein interactions in Escherichia coli. We first redesign a model tetratricopeptide repeat (TPR) protein to recognize phosphoserine in a sequence-specific fashion and characterize the interaction with its target phosphopeptide in vitro. We then combine in vivo site-specific incorporation of phosphoserine with split mCherry assembly to observe the designed phosphopeptide-protein interaction specificity in E. coli. This in vivo strategy for detecting and characterizing phosphopeptide-protein interactions has numerous potential applications for the study of natural interactions and the design of novel ones.
authors.affiliation: Department of Molecular Biophysics and Biochemistry, ‡Integrated Graduate Program in Physical and Engineering Biology, §Department of Chemistry, ∥Department of Cellular and Molecular Physiology, ⊥Systems Biology Institute, and #Department of Molecular, Cellular, and Developmental Biology, Yale University , New Haven, Connecticut 06520, United States.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Sawyer
authors.forename: Brandon M
authors.initials: BM
authors.lastname: Gassaway
authors.forename: Adrian D
authors.initials: AD
authors.lastname: Haimovich
authors.forename: Farren J
authors.initials: FJ
authors.lastname: Isaacs
authors.forename: Jesse
authors.initials: J
authors.lastname: Rinehart
authors.forename: Lynne
authors.initials: L
authors.lastname: Regan
mesh_terms: D004926:Escherichia coli / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding
chemical_list: D010750:Phosphoproteins