Valentine W M, Amarnath V, Amarnath K, Rimmele F, Graham D G
Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Chem Res Toxicol. 1995 Jan-Feb;8(1):96-102. doi: 10.1021/tx00043a013.
N,N-Diethyldithiocarbamate and its disulfide are used as pesticides, in industrial processes, and as therapeutic agents, providing numerous opportunities for human exposure. Animal studies and in vitro investigations have demonstrated adverse effects following exposure to dithiocarbamates. The ability of dithiocarbamates to decompose to parent amine and CS2 suggests that these adverse effects may be mediated through release of CS2. The toxicity of CS2 is well established, and covalent cross-linking of proteins has been presented as a potential molecular mechanism of CS2 induced neuropathy. In the present investigation the ability of N,N-diethyldithiocarbamate to effect covalent cross-linking of proteins under physiological conditions is examined. Using 13C NMR, cross-linking was observed to proceed through dithiocarbamate formation on protein amino groups followed by the production of bis(thiocarbamoyl) disulfide, dithiocarbamate ester, and thiourea cross-linking structures. The presence of bis(lysyl) thiourea cross-linking structures was verified by complete protein hydrolysis in conjunction with GC/MS. Generation of inter- and intramolecular cross-linking was established using denaturing polyacrylamide gel electrophoresis under reducing conditions and revealed that cross-linking proceeded more rapidly for N,N-diethyldithiocarbamate than for equimolar CS2 under similar conditions. Covalent cross-linking of solubilized neurofilament triplet proteins, the putative neurotoxic targets, was examined. Both N,N-diethyldithiocarbamate and CS2 were able to covalently cross-link the low molecular weight component of the neurofilament triplet proteins, but neither produced intermolecular cross-linking of the medium or high molecular weight component. These results establish that N,N-diethyldithiocarbamate promoted protein cross-linking occurs under physiological conditions and proceeds through liberation of CS2.(ABSTRACT TRUNCATED AT 250 WORDS)
N,N-二乙基二硫代氨基甲酸盐及其二硫化物被用作杀虫剂、用于工业生产过程以及作为治疗药物，这为人类接触提供了众多机会。动物研究和体外研究表明，接触二硫代氨基甲酸盐后会产生不良影响。二硫代氨基甲酸盐分解为母体胺和二硫化碳的能力表明，这些不良影响可能是通过二硫化碳的释放介导的。二硫化碳的毒性已得到充分证实，蛋白质的共价交联已被提出作为二硫化碳诱导神经病变的一种潜在分子机制。在本研究中，研究了N,N-二乙基二硫代氨基甲酸盐在生理条件下影响蛋白质共价交联的能力。使用13C核磁共振，观察到交联是通过在蛋白质氨基上形成二硫代氨基甲酸盐，随后生成双（硫代氨基甲酰基）二硫化物、二硫代氨基甲酸盐酯和硫脲交联结构来进行的。通过完全蛋白质水解结合气相色谱/质谱法验证了双（赖氨酸）硫脲交联结构的存在。在还原条件下使用变性聚丙烯酰胺凝胶电泳确定了分子间和分子内交联的产生，结果表明，在类似条件下，N,N-二乙基二硫代氨基甲酸盐的交联比等摩尔的二硫化碳进行得更快。研究了可溶性神经丝三联体蛋白（假定的神经毒性靶点）的共价交联。N,N-二乙基二硫代氨基甲酸盐和二硫化碳都能够使神经丝三联体蛋白的低分子量成分发生共价交联，但两者都不会使中分子量或高分子量成分发生分子间交联。这些结果表明，N,N-二乙基二硫代氨基甲酸盐促进的蛋白质交联在生理条件下发生，并通过二硫化碳的释放进行。（摘要截断于250字）
