Heus H A, Van Kimmenade J M, van Knippenberg P H, Hinz H J
Nucleic Acids Res. 1983 Jan 11;11(1):203-10. doi: 10.1093/nar/11.1.203.
Fragments of 16S ribosomal RNA from E. coli and B. stearothermophilus, respectively comprising the 49 and 52 3' terminal nucleotides have been studied thermodynamically using high sensitivity differential scanning calorimetry. The fragments were isolated after cleavage of 16S rRNA in the ribosome by the bacteriocin cloacin DF13. Comparison of the thermal properties of the E. coli fragments with those derived from a kasugamycin resistant mutant, which specifically lacks dimethylation of two adjacent adenosines was employed to study the effect of the methylgroups on the thermal stability. Both E. coli species exhibit similar complex melting patterns with several transitions. Overall molar transition enthalpies are equal and do not depend significantly on buffer conditions (120 kcal/mol at 15 mM Na+ to 136 kcal/mol at 215 mM Na+). However, the transition with the highest Tm, corresponding to unfolding of a nine basepair central helix is lowered by the dimethylation of the adenines in the four-membered loop. This decrease amounts to 4 degrees C at 15 mM Na+ and 2 degrees C at 215 mM Na+. The corresponding nine basepair long hairpin in the Bacillus fragment melts at a temperature of 70 degrees C at 15 mM Na+. This Tmax is much higher than expected on the basis of the sequence in the hairpin.
分别包含49个和52个3'末端核苷酸的来自大肠杆菌和嗜热栖热菌的16S核糖体RNA片段，已使用高灵敏度差示扫描量热法进行了热力学研究。这些片段是在核糖体中的16S rRNA被细菌素氯杀菌素DF13切割后分离得到的。将大肠杆菌片段的热性质与来自卡那霉素抗性突变体的片段（该突变体特别缺乏两个相邻腺苷的二甲基化）的热性质进行比较，以研究甲基对热稳定性的影响。两种大肠杆菌都表现出相似的复杂熔解模式，有多个转变。总的摩尔转变焓相等，并且在很大程度上不依赖于缓冲条件（在15 mM Na+时为120 kcal/mol，在215 mM Na+时为136 kcal/mol）。然而，对应于九碱基对中心螺旋解折叠的最高Tm的转变，因四元环中腺嘌呤的二甲基化而降低。在15 mM Na+时这种降低为4℃，在215 mM Na+时为2℃。芽孢杆菌片段中相应的九碱基对长的发夹在15 mM Na+时于70℃熔解。这个最高温度比根据发夹中的序列预期的要高得多。
