glmS ribozymes. (A) Design template and secondary structure of the glmS ribozyme used as the input for Enzymer. All of the original sequence was modified except the red nucleotides forming the catalytic core. (B) Secondary structure with a new sequence generated by Enzymer, with a color scheme corresponding to that of the design template in A. Cleavage site is shown by an arrowhead. Red nucleotides in bold are those conserved from the WT sequence. (C) Cleavage end point of the ribozymes during transcription (for 2.5 h) compared to the WT with and without glucosamine-6-P (GlcN6P). Inactivating mutations were made to the WT sequence to get the glmS uncleaved size marker, and glmS cleaved is a shorter RNA corresponding to the expected size of the cleaved product. (D) Kinetics of the ribozymes WT and glmS2 (for 64 min) compared to the WT with glucosamine-6-P (GlcN6P) and K+ or Na+.
