Tibanyenda N, De Bruin S H, Haasnoot C A, van der Marel G A, van Boom J H, Hilbers C W
Eur J Biochem. 1984 Feb 15;139(1):19-27. doi: 10.1111/j.1432-1033.1984.tb07970.x.
The stabilitye and dynamics of the duplex d(T-A-T-T-A-A--T-A-T-C-A-A-G-T-T-G) . d(C-A-A-C-T-T-G-A-T-A-T-T-A-A-T-A) has been studied by means of ultraviolet-melting, temperature-jump relaxation kinetics, stopped-flow and NMR spectroscopy. In addition, the influence of the mismatches A . A, G . T, A .C and T . C,-incorporated in this double helix through the introduction of non-complementary bases in the second strand, on these parameters has been investigated. The thermodynamic parameters characterizing the melting of the duplexes have been determined. Interestingly, a substantial decrease was observed for the values of the melting enthalpy when proceeding from 0.015 M to 0.1 M NaCl solutions. All duplexes that contain mismatches have melting temperatures below that of the totally complementary double helix. On the basis of NMR experiments and differences in the free enthalpy values between the totally complementary double helix and the duplexes with mismatches, it could be concluded that some degree of stacking of the two mispaired bases between the neighbouring base pairs is maintained. At 1 M NaCl the enthalpy and entropy of the helix-to-coil transition of the totally complementary double helix are in good agreement with values calculated on the basis of the thermodynamic data of Borer et al. [Borer, Ph. N., Dengler, B. & Tinoco, I. (1974) J. Mol. Biol. 86, 843--853] which were derived for RNA. The kinetics of the complementary duplex and duplexes with G . T and A . C mismatches were studied by means of stopped-flow and temperature-jump techniques. The rate constants of formation are the same for the three double helices. The decrease in stability of the duplexes with mismatches is therefore entirely due to an increase of the dissociation constant. In temperature-jump experiments very often a fast relaxation process is observed in addition to the relaxation characterizing the disruption of the double helix. This fast relaxation process is customarily attributed to base destacking in the single helix. By combining temperature-jump relaxation kinetics with NMR melting experiments, it is shown that at the low temperature side of the melting transition this fast relaxation process is caused by rapid changes in the double-helical structure.
通过紫外熔解、温度跃升弛豫动力学、停流和核磁共振光谱等方法，研究了双链d(T-A-T-T-A-A--T-A-T-C-A-A-G-T-T-G).d(C-A-A-C-T-T-G-A-T-A-T-T-A-A-T-A)的稳定性和动力学。此外，还研究了通过在第二条链中引入非互补碱基而掺入该双螺旋中的错配A.A、G.T、A.C和T.C对这些参数的影响。已确定了表征双链熔解的热力学参数。有趣的是，当从0.015M NaCl溶液变为0.1M NaCl溶液时，观察到熔解焓值大幅下降。所有含有错配的双链的熔解温度都低于完全互补双螺旋的熔解温度。基于核磁共振实验以及完全互补双螺旋与有错配的双链之间自由焓值的差异，可以得出结论，相邻碱基对之间两个错配碱基存在一定程度的堆积。在1M NaCl时，完全互补双螺旋的螺旋-线圈转变的焓和熵与根据Borer等人[Borer, Ph. N., Dengler, B. & Tinoco, I. (1974) J. Mol. Biol. 86, 843--853]为RNA推导的热力学数据计算得到的值高度一致。通过停流和温度跃升技术研究了互补双链以及具有G.T和A.C错配的双链的动力学。三种双螺旋的形成速率常数相同。因此，有错配的双链稳定性的降低完全是由于解离常数的增加。在温度跃升实验中，除了表征双螺旋破坏的弛豫外，经常还会观察到一个快速弛豫过程。这个快速弛豫过程通常归因于单螺旋中的碱基解堆积。通过将温度跃升弛豫动力学与核磁共振熔解实验相结合，表明在熔解转变的低温侧，这个快速弛豫过程是由双螺旋结构的快速变化引起的。
