Arakawa H, Ahmad R, Naoui M, Tajmir-Riahi H A
Department of Chemistry-Biology, University of Québec at Trois-Rivières, C.P. 500, Trois-Rivières, Québec G9A 5H7, Canada.
J Biol Chem. 2000 Apr 7;275(14):10150-3. doi: 10.1074/jbc.275.14.10150.
Chromium(VI) salts are well known to be mutagens and carcinogens and to easily cross the cell membranes. Because they are powerful oxidizing agents, Cr(VI) reacts with intracellular materials to reduce to trivalent form, which binds DNA. This study was designed to investigate the interaction of calf thymus DNA with Cr(VI) and Cr(III) in aqueous solution at pH 6.5-7.5, using Cr(VI)/DNA(P) molar ratios (r) of 1:20 to 2:1 and Cr(III)/DNA(P) molar ratios (r) of 1:80 to 1:2. UV-visible and Fourier transform infrared (FTIR) difference spectroscopic methods were used to determine the metal ion-binding sites, binding constants, and the effect of cation complexation on DNA secondary structure. Spectroscopic results showed no interaction of Cr(VI) with DNA at low anion concentrations (r = 1:20 to 1:1), whereas some perturbations of DNA bases and backbone phosphate were observed at very high Cr(VI) contents (r > 1) with overall binding constant of K = 508 M(-1). Cr(III) chelates DNA via guanine N-7 and the nearest PO(2) group with overall binding constant of K = 3.15 x 10(3) M(-1). Evidence for cation chelate formation comes from major shiftings and intensity variations of the guanine band at 1717 and the phosphate asymmetric stretching vibration at 1222 cm(-1). At low Cr(III) concentration (r = 1:40), the number of Cr(III) ions bound to DNA were 6-7 cations/500 base pairs, and this increased to 30-35 cations/500 base pairs at high metal ion content (r = 1:4). DNA condensation occurred at high cation concentration (r = 1:10). No major alteration of DNA conformation was observed, and the biopolymer remained in the B family structure upon chromium complexation.
六价铬盐是众所周知的诱变剂和致癌剂,且易于穿过细胞膜。由于它们是强氧化剂,六价铬会与细胞内物质发生反应还原为三价形式,进而与DNA结合。本研究旨在研究在pH 6.5 - 7.5的水溶液中,小牛胸腺DNA与六价铬和三价铬的相互作用,使用的六价铬/DNA(P)摩尔比(r)为1:20至2:1,三价铬/DNA(P)摩尔比(r)为1:80至1:2。采用紫外可见光谱和傅里叶变换红外(FTIR)差示光谱法来确定金属离子结合位点、结合常数以及阳离子络合对DNA二级结构的影响。光谱结果表明,在低阴离子浓度下(r = 1:20至1:1),六价铬与DNA没有相互作用,而在非常高的六价铬含量下(r > 1),观察到DNA碱基和主链磷酸基团有一些扰动,总结合常数K = 508 M⁻¹。三价铬通过鸟嘌呤N - 7和最近的PO₂基团螯合DNA,总结合常数K = 3.15×10³ M⁻¹。阳离子螯合物形成的证据来自于1717处鸟嘌呤带和1222 cm⁻¹处磷酸基团不对称伸缩振动的主要位移和强度变化。在低三价铬浓度下(r = 1:40),与DNA结合的三价铬离子数量为6 - 7个阳离子/500个碱基对,在高金属离子含量下(r = 1:4)增加到30 - 35个阳离子/500个碱基对。在高阳离子浓度下(r = 1:10)发生了DNA凝聚。未观察到DNA构象的重大改变,并且在铬络合后生物聚合物仍保持在B族结构中。