Nuclease protection analysis of ribonucleoprotein complexes: use of the cytotoxic ribonuclease alpha-sarcin to determine the binding sites for Escherichia coli ribosomal proteins L5, L18, and L25 on 5S rRNA.

摘要

A rapid and convenient method has been devised to determine the binding sites for proteins on RNA. The procedure is an adaptation of one used to map DNA-protein complexes by protection against nuclease digestion. The method uses the cytotoxic ribonuclease alpha-sarcin, which hydrolyzes purines in both single- and double-stranded regions of RNA. It has been authenticated by confirming the binding sites for the Escherichia coli ribosomal proteins L18 and L25 on 5S rRNA and its value has been established by identifying the attachment site for protein L5. The procedure should be useful for the analysis of other ribonucleoprotein complexes.