The tertiary folding of Escherichia coli 16S RNA, as studied by in situ intra-RNA cross-linking of 30S ribosomal subunits with bis-(2-chloroethyI)-methylamine

Abstract

Intra-RNA cross-links were introduced into E. coli 30S ribosomal subunits by treatment with bis-(2-chloroethyl)methylamine. The subunits were partially digested with cobra venom nuclease, and the cross-linked complexes were separated by two-dimensional electrophoresis and analysed according to our published procedures. Tertiary structural cross-links in the 16S RNA were identified between nucleotides 31 and 306, and between the tetranucleotide 693-696 and nucleotides 794 or 799. Secondary structural cross-links, lying at the ends of double-helical regions, were found between nucleotides 46 and the trinucleotide 362-364, and between the dinucleotide 148-149 and nucleotide 174. Cross-links within double-helical elements were identified between the tetranucleotide 128-131 and nucleotide 232, between nucleotide 250 and the dinucleotide 274-275, and between nucleotides 1413 and 1486. Adenine as well as guanine residues were involved in the cross-links.