title: The design and structural characterization of a synthetic pentatricopeptide repeat protein.
abstract: Proteins of the pentatricopeptide repeat (PPR) superfamily are characterized by tandem arrays of a degenerate 35-amino-acid α-hairpin motif. PPR proteins are typically single-stranded RNA-binding proteins with essential roles in organelle biogenesis, RNA editing and mRNA maturation. A modular, predictable code for sequence-specific binding of RNA by PPR proteins has recently been revealed, which opens the door to the de novo design of bespoke proteins with specific RNA targets, with widespread biotechnological potential. Here, the design and production of a synthetic PPR protein based on a consensus sequence and the determination of its crystal structure to 2.2 Å resolution are described. The crystal structure displays helical disorder, resulting in electron density representing an infinite superhelical PPR protein. A structural comparison with related tetratricopeptide repeat (TPR) proteins, and with native PPR proteins, reveals key roles for conserved residues in directing the structure and function of PPR proteins. The designed proteins have high solubility and thermal stability, and can form long tracts of PPR repeats. Thus, consensus-sequence synthetic PPR proteins could provide a suitable backbone for the design of bespoke RNA-binding proteins with the potential for high specificity.
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Benjamin S
authors: BS
authors: Gully
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Kunal R
authors: KR
authors: Shah
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Mihwa
authors: M
authors: Lee
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Kate
authors: K
authors: Shearston
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Nicole M
authors: NM
authors: Smith
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Agata
authors: A
authors: Sadowska
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Amanda J
authors: AJ
authors: Blythe
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Kalia
authors: K
authors: Bernath-Levin
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Will A
authors: WA
authors: Stanley
authors: Australian Research Council Centre of Excellence in Plant Energy Biology, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Ian D
authors: ID
authors: Small
authors: School of Chemistry and Biochemistry, The University of Western Australia, Crawley, Western Australia, Australia.
authors: Charles S
authors: CS
authors: Bond
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000737:chemistry*; D029681:Arabidopsis Proteins / Q000138:chemical synthesis / Q000737:chemistry*; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016601:RNA-Binding Proteins / Q000138:chemical synthesis / Q000737:chemistry*; D016415:Sequence Alignment
chemical_list: D029681:Arabidopsis Proteins; D016601:RNA-Binding Proteins
keywords: synthetic pentatricopeptide repeat protein