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)methylaraine. 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.