Kuykendall J R, Bogdanffy M S
Haskell Laboratory for Toxicology and Industrial Medicine, E.I. duPont de Nemours and Company, Newark, DE 19711.
Mutat Res. 1994 Nov 1;311(1):49-56. doi: 10.1016/0027-5107(94)90072-8.
The amino acid residue and nucleoside specificity of acetaldehyde-induced DNA-protein crosslinks (DPXLs) were studied using a modified filter binding assay. A 40% inhibition of acetaldehyde-induced pUC13 plasmid DNA-calf thymus histone crosslink formation was achieved by addition of 50 mM lysine (free base), while arginine was unable to affect crosslink formation at concentrations to 150 mM. Polymers (5-mers) of lysine (poly-lys5) were able to substitute for histones in acetaldehyde-induced plasmid crosslink formation, being equally effective at equimolar concentrations. Homopolymers (6-mers) of deoxyguanosine (poly-dG6) (but not deoxyadenosine, deoxycytidine or thymidine) served as an efficient substrate for acetaldehyde-induced DPXL formation, using either calf thymus histones or poly-lys5 as the protein source. Acetaldehyde-induced crosslinks between poly-dG6 and poly-lys5 were formed rapidly, but were unstable at 37 degrees C (a half-life or 1.5-2 h). Stability of these crosslinks was unaffected by pH at a range of 5.5-9.0 at 37 degrees C for 2 h. Results presented here suggest that unstable complexes of deoxyguanosine and lysine constitute a major portion of the DPXLs formed by acetaldehyde in vitro.
使用改良的滤膜结合试验研究了乙醛诱导的DNA-蛋白质交联（DPXLs）的氨基酸残基和核苷特异性。添加50 mM赖氨酸（游离碱）可使乙醛诱导的pUC13质粒DNA-小牛胸腺组蛋白交联形成受到40%的抑制，而精氨酸在浓度高达150 mM时无法影响交联形成。赖氨酸聚合物（5聚体）（聚-lys5）能够在乙醛诱导的质粒交联形成中替代组蛋白，在等摩尔浓度下同样有效。脱氧鸟苷的同聚物（6聚体）（聚-dG6）（而非脱氧腺苷、脱氧胞苷或胸腺嘧啶）可作为乙醛诱导DPXL形成的有效底物，使用小牛胸腺组蛋白或聚-lys5作为蛋白质来源。乙醛诱导的聚-dG6与聚-lys5之间的交联迅速形成，但在37℃下不稳定（半衰期为1.5 - 2小时）。在37℃下2小时，pH值在5.5 - 9.0范围内时，这些交联的稳定性不受影响。此处呈现的结果表明，脱氧鸟苷和赖氨酸的不稳定复合物构成了乙醛在体外形成的DPXLs的主要部分。
