RNA structure-dependent uncoupling of substrate recognition and cleavage by Escherichia coli ribonuclease III.
Members of the ribonuclease III superfamily of double-strand-specific endoribonucleases participate in diverse RNA maturation and decay pathways. Ribonuclease III of the gram-negative bacterium Escherichia coli processes rRNA and mRNA precursors, and its catalytic action can regulate gene expression by controlling mRNA translation and stability. It has been proposed that E.coli RNase III can function in a non-catalytic manner, by binding RNA without cleaving phosphodiesters. However, there has been no direct evidence for this mode of action. We describe here an RNA, derived from the T7 phage R1.1 RNase III substrate, that is resistant to cleavage in vitro by E.coli RNase III but retains comparable binding affinity. R1.1[CL3B] RNA is recognized by RNase III in the same manner as R1.1 RNA, as revealed by the similar inhibitory effects of a specific mutation in both substrates. Structure-probing assays and Mfold analysis indicate that R1.1[CL3B] RNA possesses a bulge- helix-bulge motif in place of the R1.1 asymmetric internal loop. The presence of both bulges is required for uncoupling. The bulge-helix-bulge motif acts as a 'catalytic' antideterminant, which is distinct from recognition antideterminants, which inhibit RNase III binding.
Calin-Jageman
Irina
I
Department of Biological Sciences, Wayne State University, Detroit, MI 48202, USA.
Nicholson
Allen W
AW
D017123:Bacteriophage T7 / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004722:Endoribonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D009843:Oligoribonucleotides / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D012313:RNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D043244:Ribonuclease III; D013379:Substrate Specificity
D029968:Escherichia coli Proteins; D009843:Oligoribonucleotides; D012367:RNA, Viral; D012313:RNA; D004722:Endoribonucleases; D043244:Ribonuclease III; C468366:ribonuclease III, E coli