title: Analog sensitive chemical inhibition of the DEAD-box protein DDX3.
abstract: Proper maintenance of RNA structure and dynamics is essential to maintain cellular health. Multiple families of RNA chaperones exist in cells to modulate RNA structure, RNA-protein complexes, and RNA granules. The largest of these families is the DEAD-box proteins, named after their catalytic Asp-Glu-Ala-Asp motif. The human DEAD-box protein DDX3 is implicated in diverse biological processes including translation initiation and is mutated in numerous cancers. Like many DEAD-box proteins, DDX3 is essential to cellular health and exhibits dosage sensitivity, such that both decreases and increases in protein levels can be lethal. Therefore, chemical inhibition would be an ideal tool to probe the function of DDX3. However, most DEAD-box protein active sites are extremely similar, complicating the design of specific inhibitors. Here, we show that a chemical genetic approach best characterized in protein kinases, known as analog-sensitive chemical inhibition, is viable for DDX3 and possibly other DEAD-box proteins. We present an expanded active-site mutant that is tolerated in vitro and in vivo, and is sensitive to chemical inhibition by a novel bulky inhibitor. Our results highlight a course towards analog sensitive chemical inhibition of DDX3 and potentially the entire DEAD-box protein family.
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, California.|Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, California.
authors.forename: Stephen N
authors.initials: SN
authors.lastname: Floor
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California.
authors.forename: Krister J
authors.initials: KJ
authors.lastname: Barkovich
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, California.
authors.forename: Kendall J
authors.initials: KJ
authors.lastname: Condon
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California.|Howard Hughes Medical Institute, University of California, San Francisco, California.|Department of Chemistry, University of California, Berkeley, California.
authors.forename: Kevan M
authors.initials: KM
authors.lastname: Shokat
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, California.|Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, California.|Department of Chemistry, University of California, Berkeley, California.|Innovative Genomics Initiative, University of California, Berkeley, Berkeley, California.|Lawrence Berkeley National Laboratory, Physical Biosciences Division, Berkeley, California.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Doudna
mesh_terms: D000249:Adenosine Monophosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D020134:Catalytic Domain / Q000187:drug effects; D053487:DEAD-box RNA Helicases / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D009154:Mutation; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D013844:Thiazoles / Q000737:chemistry* / Q000494:pharmacology*
chemical_list: D013844:Thiazoles; D000249:Adenosine Monophosphate; C027348:2-anilino-5-thiazolinone; D012313:RNA; C119332:DDX3X protein, human; D053487:DEAD-box RNA Helicases
keywords: DDX3 inhibitor; DEAD-box proteins; RNA; chemical genetics; protein engineering; small-molecule inhibitor