Structural basis for binding of the plasmid ColIb-P9 antisense Inc RNA to its target RNA with the 5'-rUUGGCG-3' motif in the loop sequence.
The sequence 5'-rUUGGCG-3' is conserved within the loop regions of antisense RNAs or their targets involved in replication of various prokaryotic plasmids. In IncIalpha plasmid ColIb-P9, the partially base paired 21-nucleotide loop of a stem-loop called structure I within RepZ mRNA contains this hexanucleotide sequence, and comprises the target site for the antisense Inc RNA. In this report, we find that the base pairing interaction at the 5'-rGGC-3' sequence in the hexanucleotide motif is important for interaction between Inc RNA and structure I. In addition, the 21-base loop domain of structure I is folded tighter than predicted, with the hexanucleotide sequence at the top. The second U residue in the sequence is favored for Inc RNA binding in a base-specific manner. On the other hand, the upper domain of the Inc RNA stem-loop is loosely structured, and maintaining the loop sequence single-stranded is important for the intermolecular interaction. Based on these results, we propose that a structural feature in the loop I domain, conferred probably by the conserved 5'-rUUGGCG-3' sequence, favors binding to a complementary, single-stranded RNA. This model also explains how the RepZ mRNA pseudoknot, described in the accompanying paper (Asano, K., and Mizobuchi, K. (1998) J. Biol. Chem. 273, 11815-11825) is formed specifically with structure I. A possible conformation adopted by the 5'-rUUGGCG-3' loop sequence is discussed.
Asano
K
K
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Hongo, Tokyo 113, Japan.
Niimi
T
T
Yokoyama
S
S
Mizobuchi
K
K
D001426:Bacterial Proteins / Q000235:genetics; D003086:Bacteriocin Plasmids; D001483:Base Sequence; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D006860:Hydrogen Bonding; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D016372:RNA, Antisense / Q000235:genetics*; D012329:RNA, Bacterial / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D016372:RNA, Antisense; D012329:RNA, Bacterial; D012333:RNA, Messenger; C112255:RepZ protein, plasmid ColIb-P9