THE ALKALINE DENATURATION OF DEOXYRIBOSE NUCLEIC ACID
THE ALKALINE DENATURATION OF DEOXYRIBOSE NUCLEIC ACID
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DOI：
10.1021/ja01549a033
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发表时间：
1958-01-01
影响因子：
15
通讯作者：
DOTY, P
中科院分区：
文献类型：
--
作者：
EHRLICH, P;DOTY, P
采用沉淀法、黏度法和光散射法对小牛胸腺脱氧核糖核酸（DNA）在pH值7 ~ 12.5范围内进行了检测。当离子强度为0.15时，发现在pH 11.7附近分子构型发生急剧转变。pH值高于11.9时，转变完成，DNA保持原来的分子量，但体积缩小了。特性粘度降低了10倍，在259µ时光密度增加了12%，旋转半径减小了近3倍，平均沉降常数不变，这与之前在酸变性（pH 3.1）和热脱自然中观察到的变化基本相同。当离子强度降低到0.005时，碱性变性DNA发生了非常大的分子膨胀。在相同条件下，光散射结果表明，天然DNA的结构保持不变。所有这些观察结果都与以下观点一致：未变性的DNA是高度延伸和相对刚性的，因为沃森-克里克结构的两条多核苷酸链之间存在连续的氢键，而变性状态由随机卷曲的、高度灵活的链组成，由于大量非周期性排列的分子内氢键的持续存在，这些链保持成对和高度收缩。1947年Gulland， Jordan和Taylor1表明，当脱氧核糖核酸（DNA）溶液酸化或碱性时，其粘度急剧下降。对酸2·3的影响的研究表明，在0.2离子强度下小心地向下调整pH，在pH 3.1附近发生急剧转变，到pH 2.6时，DNA已经达到了一个新的、相对稳定的状态。另一项研究表明，在相同的离子强度下，在90度附近加热时发生了类似的转变。在这两种情况下，构型转变的标志都是在恒定分子量下非常显著的分子收缩。这被解释为周期性氢键的破裂，DNA的沃森-克里克结构形成随机卷曲和更紧密的状态，残余的分子内氢键使两个多核苷酸链连接在一起。这里报道的调查是为了完成DNA变性的研究。我们特别想看看它是否遵循酸性和热变性建立的相同模式，看看碱性条件是否可能导致被认为构成天然DNA分子的两条多核苷酸链的分离，并检查离子强度降低对天然和未变性形式的分子构型的影响。
Deoxyribose nucleic acid (DNA) from calf thymus has been examined by sedimentation, viscosity and light scattering methods in the pH range from 7 to 12.5. When the ionic strength is 0.15, it is found that the molecular configuration under-goes a sharp transition in the vicinity of pH 11.7. Above pH 11.9 the transition is complete and the DNA is found to re-tain its original molecular weight but to have contracted in size. The 10-fold lowering of the intrinsic viscosity, the12% rise inoptical density at 259 µ, the nearly 3-fold reduction in radius of gyration and the invariance of the mean sedimentation constant are essentially identical with the changes previously observed for acid denaturation (pH 3.1) andthermal de-naturation. Alkaline denatured DNA undergoes a very large molecular expansion when the ionic strength is lowered to 0.005. Under the same conditions light scattering results show that the configuration of native DNA remains unchanged. All of these observations are consistent with the view that the undenatured DNA is highly extended and relatively rigid because of thecontinuous hydrogen bonding between the two polynucleotide chains of the Watson-Crick structure, whereas the denatured state consists of randomly coiled, highly flexible chains that remain paired and highly contracted dueto the persistence of a substantial number of non-periodically arranged, intramolecular hydrogen bonds.In 1947 Gulland, Jordan and Taylor1 showed that the viscosity of deoxyribose nucleic acid (DNA) solutions fell sharply when the solutions were acidified or made alkaline. Astudy of the effects of acid2· 3 demonstrated that upon careful downward adjustment of the pH at 0.2 ionic strength, a sharp transition occurred in the vicinity of pH 3.1 and that by pH 2.6 a new, relatively stable state of DNA had been attained. Another study4showed that at the same ionic strength a comparable transition occurred upon heating in the vicinity of 90. In both cases the configurational transition was marked by very substantial molecular con-traction at constant molecular weight. This was interpreted as a breakdown of the periodically hy-drogen bonded, Watson-Crick structure of DNA to form a randomly coiled and much more compact state in which the residual intramolecular hydrogen bonds kept thetwo polynucleotide chains united. The investigation reported here was undertaken to complete this study of the denaturation of DNA. In particular we wished to see if it followed the same pattern established for acidic and thermal denaturation, to see if the alkaline conditions might lead to a separation of the two polynucleotide chains thought to make up the native DNA molecule and to examine the effects of diminished ionic strength on the molecular configuration of both the native and undenatured forms.