title: Auto-inhibition of the Dbl family protein Tim by an N-terminal helical motif.
abstract: Dbl-related oncoproteins are guanine nucleotide exchange factors specific for Rho-family GTPases and typically possess tandem Dbl homology (DH) and pleckstrin homology domains that act in concert to catalyze exchange. Because the ability of many Dbl-family proteins to catalyze exchange is constitutively activated by truncations N-terminal to their DH domains, it has been proposed that the activity of Dbl-family proteins is regulated by auto-inhibition. However, the exact mechanisms of regulation of Dbl-family proteins remain poorly understood. Here we show that the Dbl-family protein, Tim, is auto-inhibited by a short, helical motif immediately N-terminal to its DH domain, which directly occludes the catalytic surface of the DH domain to prevent GTPase activation. Similar to the distantly related Vav isozymes, auto-inhibition of Tim is relieved by truncation, mutation, or phosphorylation of the auto-inhibitory helix. A peptide comprising the helical motif inhibits the exchange activity of Tim in vitro. Furthermore, substitutions within the most highly conserved surface of the DH domain designed to disrupt interactions with the auto-inhibitory helix also activate the exchange process.
authors.affiliation: Department of Pharmacology, University of North Carolina, Chapel Hill, North Carolina 27599-7295, USA.
authors.forename: Marielle E
authors.initials: ME
authors.lastname: Yohe
authors.forename: Kent L
authors.initials: KL
authors.lastname: Rossman
authors.forename: Olivia S
authors.initials: OS
authors.lastname: Gardner
authors.forename: Antoine E
authors.initials: AE
authors.lastname: Karnoub
authors.forename: Jason T
authors.initials: JT
authors.lastname: Snyder
authors.forename: Svetlana
authors.initials: S
authors.lastname: Gershburg
authors.forename: Lee M
authors.initials: LM
authors.lastname: Graves
authors.forename: Channing J
authors.initials: CJ
authors.lastname: Der
authors.forename: John
authors.initials: J
authors.lastname: Sondek
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics; D000818:Animals; D002384:Catalysis; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation / Q000235:genetics; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009154:Mutation; D017434:Protein Structure, Tertiary / Q000235:genetics; D051567:Proto-Oncogene Proteins c-vav / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D020662:Guanine Nucleotide Exchange Factors; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; C512843:Mcf2 protein, mouse; D051567:Proto-Oncogene Proteins c-vav; D011994:Recombinant Proteins; C509769:TIMELESS protein, human; C509770:Timeless protein, mouse