title: Arginine arrangement of bacteriophage λ N-peptide plays a role as a core motif in GNRA tetraloop RNA binding.
abstract: A simple α-helical N-model-peptide was designed to investigate the role of the arginine-rich motif of bacteriophage λ N-peptide in selective binding with boxB RNA. The five-arginine arrangement of native N-peptide was retained; all other residues were replaced with alanine. In vitro selection of RNA (30 random-nucleotide region) was carried out with N-model-peptide immobilized on a 27 MHz quartz-crystal microbalance (QCM). Selected RNAs were evaluated on the same QCM plate to obtain binding constants (Ka =10(7) -10(8)  M(-1) ). Many selected RNAs contained GNR(N)A-type loops (similar to the boxB RNA motif recognized by the native N-peptide). Fragments and minimal RNAs containing the GNRA-type loop also bound to N-model-peptide (Ka =10(6) -10(7)  M(-1) ). The RNA recognition specificity of the peptide was studied by changing the "closing" U-A base pair and one base in the tetraloop of the RNA aptamers, and by peptide mutations (18th residue of N-model-peptide). It was concluded that the five-arginine arrangement of the peptide performs selective recognition of the GNRA tetraloop and GNR(N)A pentaloop RNA structures, and that substitution of another functional amino acid residue at the 18th position in N-peptide adds the recognition ability for a loop-RNA sequence.
authors.affiliation: Innovative Flex Course for Frontier Organic Material Systems (iFront), Yamagata University, 4-3-16 Jonan, Yonezawa, Yamagata 992-8510 (Japan); Department of Biomolecular Engineering, Tokyo Institute of Technology, 4259, Nagatsuda, Midori-ku, Yokohama, 226-8501 (Japan).
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Furusawa
authors.forename: Shinobu
authors.initials: S
authors.lastname: Fukusho
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Okahata
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000378:metabolism; D001120:Arginine / Q000737:chemistry*; D001435:Bacteriophages / Q000378:metabolism*; D020029:Base Pairing; D055767:Immobilized Proteins / Q000737:chemistry / Q000378:metabolism; D009690:Nucleic Acid Conformation; D059372:Nucleotide Motifs; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D058525:Quartz Crystal Microbalance Techniques; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism*
chemical_list: D052157:Aptamers, Nucleotide; D055767:Immobilized Proteins; D010455:Peptides; D012367:RNA, Viral; D001120:Arginine
keywords: N-peptides; QCM; RNA structures; helical structures; in vitro selection; peptides