Zhitkovich A, Voitkun V, Costa M
Nelson Institute of Environmental Medicine, New York University Medical Center, New York 10016, USA.
Biochemistry. 1996 Jun 4;35(22):7275-82. doi: 10.1021/bi960147w.
We have recently shown that a substantial fraction of all Cr-DNA adducts in chromate-exposed cells are represented by ternary complexes involving amino acids or glutathione bridged by Cr-(III) to DNA. The tridentate amino acids such as cysteine, glutamic acid, and histidine were predominantly found cross-linked to DNA. The mechanism by which Cr can cross-link these amino acids to DNA has been modeled by reacting DNA and trivalent and hexavalent chromium with cysteine and histidine. The formation of a Cr(III)-amino acid binary complex was required before Cr(III) reacted with DNA to yield a ternary complex. Cr(III)-pretreated DNA did not bind cysteine or histidine even after prolonged incubations. Reduction of Cr(VI) in the presence of DNA gave rise to an extensive cross-linking of cysteine and histidine. Addition of DNA to Cr(VI) mixtures at the start of reduction or after the reduction was complete had little effect on the level of ternary complexes indicating that Cr(III)-amino acid binary complexes were DNA-attacking species. In order to identify DNA groups involved in the ternary complex formation, pre-formed Cr(III)-histidine complexes were reacted with nucleosides and nucleotide monophosphates followed by separation and analysis of the products. The incubation of the Cr(III)-histidine complexes with nucleotide monophosphates but not with nucleosides gave rise to ternary complexes that contained both histidine and Cr, showing the primary importance of the phosphate group in this reaction. All four DNA nucleotides were capable of the ternary complex formation with Cr(III) and histidine. No apparent base preference in the amino acid cross-linking was also found in the reaction of Cr(III)/cysteine and Cr(VI)/cysteine mixtures with oligonucleotides of base-specific composition.
我们最近发现，在暴露于铬酸盐的细胞中，所有铬 - DNA加合物的很大一部分是由三元复合物构成的，这些复合物涉及通过Cr(III)与DNA桥接的氨基酸或谷胱甘肽。主要发现三齿氨基酸如半胱氨酸、谷氨酸和组氨酸与DNA交联。通过使DNA以及三价和六价铬与半胱氨酸和组氨酸反应，模拟了Cr将这些氨基酸交联到DNA的机制。在Cr(III)与DNA反应生成三元复合物之前，需要先形成Cr(III)-氨基酸二元复合物。即使长时间孵育，经Cr(III)预处理的DNA也不结合半胱氨酸或组氨酸。在DNA存在下还原Cr(VI)会导致半胱氨酸和组氨酸广泛交联。在还原开始时或还原完成后向Cr(VI)混合物中添加DNA对三元复合物水平影响很小，这表明Cr(III)-氨基酸二元复合物是攻击DNA的物种。为了鉴定参与三元复合物形成的DNA基团，将预先形成的Cr(III)-组氨酸复合物与核苷和核苷酸单磷酸反应，然后分离并分析产物。Cr(III)-组氨酸复合物与核苷酸单磷酸而非核苷孵育会产生同时含有组氨酸和Cr的三元复合物，这表明磷酸基团在该反应中至关重要。所有四种DNA核苷酸都能够与Cr(III)和组氨酸形成三元复合物。在Cr(III)/半胱氨酸和Cr(VI)/半胱氨酸混合物与具有碱基特异性组成的寡核苷酸的反应中，也未发现氨基酸交联中有明显的碱基偏好。
