Takeshita M, Kappen L S, Grollman A P, Eisenberg M, Goldberg I H
Biochemistry. 1981 Dec 22;20(26):7599-606. doi: 10.1021/bi00529a039.
The nucleotide sequence specificity of neocarzinostatin (NCS), auromomycin (AUR), bleomycin (Blm), phleomycin (Phlm), and tallysomycin (Tlm) has been determined by using these antibiotics and their associated chromophores to create strand scissions in end-labeled restriction fragments of DNA and then determining the base sequence of the oligonucleotides formed. NCS and the NCS chromophore induce similar patterns of cleavage in DNA fragments labeled at the 5' terminus. The pattern produced by the AUR chromophore also resembles that of its holoantibiotic. Dithiothreitol enhances the rate of cleavage of DNA by the AUR chromophore but does not alter the sequence specificity. The results suggest that the polypeptide component of AUR and NCS serves primarily as a carrier for the chromophore. When tested with a fragment labeled at the 3' terminus, the products of NCS and AUR cleavage do not display the patterns of chemically produced oligonucleotides cleaved at phosphodiester bonds, suggesting that the 5' terminus is modified by a sugar fragment. NCS primarily attacks thymine (75% of the total bases attacked) and, to a lesser extent, adenine (19%) and cytosine (6%). AUR preferentially attacks guanine (67% of total bases), while attacking less often thymine (24%) and adenine (9%). Bleomycin and its analogues preferentially cleave purine--pyrimidine (5' leads to 3') and pyrimidine--pyrimidine (3' leads to 5') sequences. All (5' leads to 3') GT and GC sequences were cleaved. Phlm G and Phlm-Pep are less active than bleomycin toward purines while Tlm was more active. The patterns of cleavage produced by Blm A2 and Blm B6 are similar, while those produced by Phlm-Pep, Phlm G, Blm-B1', and Blm-Pep resemble one another. The cleavage pattern of Tlm shows quantitative differences from the other analogues tested. Differences between bleomycin and its analogues may be related to structural differences in these molecules.
通过使用新制癌菌素（NCS）、金霉素（AUR）、博来霉素（Blm）、腐草霉素（Phlm）和 tallysomycin（Tlm）这些抗生素及其相关发色团，在末端标记的 DNA 限制片段中产生链断裂，然后确定形成的寡核苷酸的碱基序列，已确定了它们的核苷酸序列特异性。NCS 及其发色团在 5' 末端标记的 DNA 片段中诱导相似的切割模式。AUR 发色团产生的模式也与其全抗生素的模式相似。二硫苏糖醇提高了 AUR 发色团切割 DNA 的速率，但不改变序列特异性。结果表明，AUR 和 NCS 的多肽成分主要作为发色团的载体。当用 3' 末端标记的片段进行测试时，NCS 和 AUR 切割的产物未显示出在磷酸二酯键处化学产生的寡核苷酸的切割模式，这表明 5' 末端被一个糖片段修饰。NCS 主要攻击胸腺嘧啶（占总攻击碱基的 75%），在较小程度上攻击腺嘌呤（19%）和胞嘧啶（6%）。AUR 优先攻击鸟嘌呤（占总碱基的 67%），而较少攻击胸腺嘧啶（24%）和腺嘌呤（9%）。博来霉素及其类似物优先切割嘌呤 - 嘧啶（5' 至 3'）和嘧啶 - 嘧啶（3' 至 5'）序列。所有（5' 至 3'）的 GT 和 GC 序列均被切割。Phlm G 和 Phlm - Pep 对嘌呤的活性低于博来霉素，而 Tlm 更具活性。Blm A2 和 Blm B6 产生的切割模式相似，而 Phlm - Pep、Phlm G、Blm - B1' 和 Blm - Pep 产生的模式彼此相似。Tlm 的切割模式与测试的其他类似物显示出定量差异。博来霉素及其类似物之间的差异可能与这些分子的结构差异有关。
