Surovaia A N, Grokhovskiĭ S L, Brusov R V, Lysov Iu P, Zhuze A L, Gurskiĭ G V
Mol Biol (Mosk). 1994 Nov-Dec;28(6):1383-99.
De novo design and synthesis by a solid phase technique of linear and cyclic 26-residues peptides are reported. The peptides use beta-strand-turn-beta -strand motif for sequence recognition on DNA. Amino acid sequences in the two peptides are identical, but the structure of the cyclic peptide is constrained by S-S bridge between two cysteine residues. A 28-residue peptide containing at the N-terminus a copper-chelating peptide Gly-Gly-His is also synthesized which can be used as a potential DNA-cleaving reagent. Binding of these peptides to various natural and synthetic DNAs and DNA fragment with a known base pair sequence has been studied by CD spectroscopy, fluorescence methods and DNAse I footprinting technique. By means of CD spectroscopy it is shown that 26-residue linear and cyclic peptides are partially in disordered and beta-conformations in aqueous solution in absence and in presence of 20% trifluoroethanol (TFE), but assume partially an alpha-helix conformation in the presence of 50% TFE. It is shown that linear and cyclic peptides bind to DNA. The binding approaches saturation level when one peptide molecule is bound approximately per three or four DNA base pairs. We found that antibiotic distamycin A, binding in the minor DNA groove, competes effectively with the 26-residue linear and cyclic peptides for binding to poly(dA).poly (dT). According to the CD spectroscopy data the linear and cyclic peptides undergo conformation changes upon binding to DNA, whereas the DNA structure is not markedly altered. Difference CD spectra obtained by subtracting the spectrum of the free DNA from the spectrum of the peptide-DNA mixture differ from the spectrum of the free peptide. The shapes of difference CD spectra are consistent with a conformation transition from a disordered conformation into a beta-like conformation upon binding of peptide to DNA. DNAase I footprinting diagrams show that there is a specific protection by linear and cyclic peptides of the nucleotide sequences on two ends of operators OR1, OR2 and OR3 and pseudooperators within the cro gene of 434 phage.
报道了通过固相技术从头设计并合成线性和环状26残基肽。这些肽利用β-链-转角-β-链基序在DNA上进行序列识别。两种肽的氨基酸序列相同，但环状肽的结构由两个半胱氨酸残基之间的S-S桥所限制。还合成了一种28残基肽，其N端含有铜螯合肽Gly-Gly-His，可作为潜在的DNA切割试剂。通过圆二色光谱（CD光谱）、荧光方法和DNA酶I足迹技术研究了这些肽与各种天然和合成DNA以及具有已知碱基对序列的DNA片段的结合。通过CD光谱表明，26残基线性和环状肽在不存在和存在20%三氟乙醇（TFE）的水溶液中部分处于无序和β构象，但在存在50% TFE时部分呈现α-螺旋构象。结果表明线性和环状肽与DNA结合。当每三或四个DNA碱基对大约结合一个肽分子时，结合接近饱和水平。我们发现，结合于DNA小沟的抗生素偏端霉素A与26残基线性和环状肽有效竞争结合聚（dA）·聚（dT）。根据CD光谱数据，线性和环状肽在与DNA结合时会发生构象变化，而DNA结构没有明显改变。通过从肽-DNA混合物的光谱中减去游离DNA的光谱获得的差示CD光谱不同于游离肽的光谱。差示CD光谱的形状与肽与DNA结合时从无序构象转变为β样构象的构象转变一致。DNA酶I足迹图表明，线性和环状肽对434噬菌体cro基因内的操纵子OR1、OR2和OR3两端的核苷酸序列以及假操纵子有特异性保护作用。
