Vardimon L, Rich A
Proc Natl Acad Sci U S A. 1984 Jun;81(11):3268-72. doi: 10.1073/pnas.81.11.3268.
Plasmids carrying 24- or 32-base-pair inserts of alternating (dG-dC) residues were used to analyze the level of methylation of the G-C-G-C sites by Hha I DNA methyltransferase and their cleavage by Hha I endonuclease in the B-DNA or Z-DNA conformation. In supercoiled plasmids in which the inserts formed Z-DNA, the extent of methylation at the insert G-C-G-C sites was dramatically lower than the level of methylation at the G-C-G-C sites located outside the insert in the same plasmid. Similarly, cleavage by Hha I endonuclease was sharply lowered when the insert was in the Z-DNA form. In the relaxed plasmid, all its G-C-G-C sites were methylated to the same extent and the unmethylated sites were readily cleaved. After treatment with the methylase, the supercoiled plasmid was linearized and then digested with Hha I restriction endonuclease. This exposed unmethylated G-C-G-C sites from the insert that had been protected against cleavage in the Z conformation. A chemical reaction was used to study the distribution of the unmethylated cytosine residues. No accumulation of unmethylated cytosine residues was found anywhere along the entire 32-base-pair insert, which is consistent with a cooperative B-Z transition.
携带由交替的（dG-dC）残基组成的24或32个碱基对插入片段的质粒，被用于分析Hha I DNA甲基转移酶对G-C-G-C位点的甲基化水平，以及在B-DNA或Z-DNA构象中Hha I核酸内切酶对其的切割情况。在插入片段形成Z-DNA的超螺旋质粒中，插入片段G-C-G-C位点的甲基化程度显著低于同一质粒中位于插入片段之外的G-C-G-C位点的甲基化水平。同样，当插入片段呈Z-DNA形式时，Hha I核酸内切酶的切割也会急剧降低。在松弛质粒中，其所有G-C-G-C位点的甲基化程度相同，未甲基化的位点很容易被切割。用甲基化酶处理后，超螺旋质粒被线性化，然后用Hha I限制性核酸内切酶消化。这暴露了来自插入片段的未甲基化的G-C-G-C位点，这些位点在Z构象中曾受到保护而不被切割。利用化学反应研究未甲基化胞嘧啶残基的分布。在整个32个碱基对的插入片段上，未发现未甲基化胞嘧啶残基的积累，这与协同的B-Z转变一致。
