Engineering RNA sequence specificity of Pumilio repeats.
Puf proteins bind RNA sequence specifically and regulate translation and stability of target mRNAs. A "code" for RNA recognition has been deduced from crystal structures of the Puf protein, human Pumilio1, where each of eight repeats binds an RNA base via a combination of three side chains at conserved positions. Here, we report the creation of seven soluble mutant proteins with predictably altered sequence specificity, including one that binds tightly to adenosine-uracil-rich element RNA. These data show that Pumilio1 can be used as a scaffold to engineer RNA-binding proteins with designed sequence specificity.
Cheong
Cheom-Gil
CG
Laboratory of Structural Biology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709, USA.
Hall
Traci M Tanaka
TM
D000225:Adenine / Q000737:chemistry; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D015202:Protein Engineering; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid / Q000235:genetics; D014498:Uracil / Q000737:chemistry
C471242:PUM1 protein, human; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D014498:Uracil; D000225:Adenine