MIYUN KWON and SCOTT A. STROBEL
In the above-mentioned article, the authors reported that the P1 helix of aptamer 1 and the P3a helices of aptamers 1 and 2 are important for cooperative glycine binding based upon in-line probing results of RNAs containing point mutations in these regions (see data in Fig. 7 and Table 1). Several of these point mutations were reanalyzed for cooperativity as part of a follow-up study. Hill coefficients for mutations in the P1 and P3a regions of aptamer 1 (U12-G127, G67-U77 and G67-C77) were in agreement with those reported, consistent with a role for these helices in cooperative ligand binding. However, we were unable to reproduce the anti-cooperative effect (Hill coefficient of 0.6) reported for the G174-U206 mutation in the P3b region of aptamer 2. Reanalysis of this mutant produced a Hill coefficient of 1.5 ± 0.2, which is the same value as wild type. Thus while the data support the conclusion that the P3a region of aptamer 2 is important for glycine binding (based upon in-line probing in Fig. 2 and nucleotide analog interference mapping in Fig. 5), the revised data do not support a role for this helix in mediating a direct cooperative interaction between the two aptamers.