The three-dimensional folding of ribosomal RNA; localization of a series of intra-RNA cross-links in 23S RNA induced by treatment ofEscherichia coli50S ribosomal subunits withbis-(2-chloroethyl)-methylamine

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Abstract
Intact 50S ribosomal subunits from E.coli were cross-linked with the symmetrical bifunctlonai reagent bis (2-chloroethyl)-methylam ne. After deproteinization, selected regions of the 23S RNA were excised by treatment with ribonuclease H in the presence of appropriate complementary decadeoxynucleotides, and screened for the presence of intra-RNA cross-links by two-dimensional gel electrophoresis. Individual isolated cross-linked RNA fragments were analysed by our established procedures. Sixteen intra-RNA cross-links were identified, three of which corresponded to those previously published. The thirteen ‘new’ cross-links were localized in the 23S RNA at positions 774–78 linked to 792–94, 876–79 linked to 899–900, 979–81 or 983–84 to 2029, 1715 to 1743–46, 1911–21 to 1964, 1933 to 1966, 2032 to 2054–55, 2112 to 2169–71, 2116–17 to 2163–67, 2128–32 to 2156–59, 2392–93 to 2422–23, 2737–38 to 2763–66, and 2791 to 2890. These results are discussed in the context of three-dimensional model-building studies with the 23S RNA, with particular reference to the environment of the ‘active centre’ of the 50S subunit.

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