Escherichia coli 5S RNA A and B conformers. Characterisation by enzymatic and chemical methods

The structures of the two stable conformers of Escherichia coli 5 S RNA, the and B form, were compared. Information about the structures were obtained using the methods of limited enzymatic hydrolysis and chemical modification of accessible nucleotides. Base-specific modifications were performed for adenosines and cytidines using diethylpyrocarbonate and dimethylsulfate in combination with a strand-scission reaction at the modified site. Base-specific (RNase T1) as well as conformation-specific (nuclease S1, cobra venom nuclease) enzymes were employed for the limited enzymatic hydrolysis. Clear differences in the accessibility of the two 5 S RNA conformers to the enzymes and the chemical reagents were established and the regions with altered reactivities were localized in the 5 S RNA structure. The results are consistent with the disruption of the secondary structural interactions in helix II and partly in helices III and IV during the transition from the A to the B form. (The numbering of the helices is according to the generally accepted Fox and Woese model.) In addition some regions presumably involved in the tertiary structure are distorted. There is evidence, however, for the new formation of structural regions between two distant sites in the 5 S RNA B form. The results enable us to refine the existing 5 S RNA A-form model and provide insight into the structural dynamics that lead to the formation of the 5 S RNA B form.