FIGURE 6.

Enzymatic probing of RAAA substitution mutant. (A) RNase T1 probing analyzed by primer extension. RNA harboring the CGCCC substitution in the RAAA motif was incubated with RNase T1. A filled arrow denotes changes with the pattern obtained in the wild-type domain 3 (Fernández-Miragall and Martínez-Salas 2003). (B) Direct RNase T1 cleavage pattern of [γ-³²P]ATP-5′-end-labeled CGCCC RNA, in comparison to the wild-type (AAAAG) RNA. Bands with the same electrophoretic mobility as cleavage fragments of the untreated transcript were not considered. Closed and empty arrows are used for new and lack of attack, respectively, relative to the wild-type sequence. (C) Reorganization of RNA structure in CGCCC mutant. Results from DMS attack have been represented on the secondary structure derived from RNase A, T1, and V1 digestion.