Figure 2.

Miscleavage of various substrates. (A) Identification of the cleavage site in pGlnU/A by primer extension analysis as described in Materials and Methods. In this experiment, ∼0.2 μM pGlnU/A was incubated for 8 min (after preincubation of M1 RNA for 7 min) in the presence or absence of 2 μM M1 RNA in 50 mM MES (pH 6.0) and 40 mM Mg(OAc)₂. This was followed by primer extension analysis using an oligodeoxynucleotide complementary to residues A31 through C48 in tRNA^Gln (see Figure 1). Cs = cleavage site. (B and C) Miscleavage of various pATSer substrates [(B) with C at −1 and (C) with U at −1) represented as percentage of miscleavage and cleavage sites as indicated where for example UGG/C corresponds to the substrate pATSerUGG/C, for details see text. The 5′ cleavage fragments were separated on 22% denaturing PAGE as described in Materials and Methods.