Involvement of precursor-specific segments in the in vitro maturation of Bacillus subtilis precursor 5S ribosomal RNA

Abstract

In vitro maturation of precursor 5S ribosomal RNA (p5A) from B. subtilis effected by RNase M5 yields mature 5S RNA (m5, 116 nucleotides), and 3'' precursor-specific segment (42 nucleotides), and a 5'' precursor-specific segment (21 nucleotides). Limited digestion of p5A with RNase T2 introduces a single scission at position 60 of the molecule; m5 is cleaved at the corresponding nucleotide residue. The complementary halves of the molecules could be isolated from denaturing polyacrylamide gels. The isolated fragments of p5A are not substrates for RNase M5, suggesting that some recognition elements can be utilized by RNase M5 only when presented in double-helical form. In exploring the involvement of the precursor-specific segments in the RNase M5-p5A interaction, substrate molecules lacking the 3'' or 5'' precursor-specific segment were constructed by reannealing complementary halves from p5A and m5 RNA. The artificial substrate lacking the 5''-terminal precursor segment was cleaved very much more slowly than that lacking t'' segment; the 5'' precursor-specific segment therefore contains one or more components recognized by RNase M5 during its interaction with the p5A substrate.