INHIBITION OF 2,5-HEXANEDIONE-INDUCED PROTEIN CROSS-LINKING BY BIOLOGICAL THIOLS - CHEMICAL MECHANISMS AND TOXICOLOGICAL IMPLICATIONS

被引:21
作者
ZHU, MS
SPINK, DC
YAN, B
BANK, S
DECAPRIO, AP
机构
[1] NEW YORK STATE DEPT HLTH,WADSWORTH CTR LABS & RES,HUMAN TOXICOL & MOLEC EPIDEMIOL LAB,ALBANY,NY 12201
[2] SUNY ALBANY,DEPT ENVIRONM HLTH & TOXICOL,ALBANY,CA
[3] SUNY ALBANY,DEPT CHEM,ALBANY,CA
关键词
D O I
10.1021/tx00047a017
中图分类号
R914 [药物化学];
学科分类号
100701 ;
摘要
n-Hexane is metabolized to the gamma-diketone 2,5-hexanedione (2,5-HD), a derivative that covalently binds to lysine residues in neurofilament (NF) protein to yield 2,5-dimethylpyrrole adducts. Studies comparing the pyrrole-forming potential and neurotoxic potency of gamma-diketones have demonstrated that pyrrolylation is an absolute requirement in the neuropathogenesis. Autoxidative cross-linking of pyrrolylated NF proteins occurs and is proposed as a second required event. In the present study, the role of nucleophilic thiols and amines in the pyrrole-mediated cross-linking reaction was investigated. When pyrrolylated ribonuclease was incubated with N-acetyllysine, N-acetylcysteine, or glutathione in physiologic buffer (pH 7.4) under air, pyrrole-to-pyrrole cross-linking was inhibited only by the thiol-containing compounds. Stable thiol-pyrrole conjugates containing a bridge from the pyrrole ring at C-3 to the sulfur atom of the thiol were characterized by thermospray LC/MS and H-1-NMR spectroscopy. In contrast to low-molecular-mass thiols, SDS-PAGE studies indicated that, under the same incubation conditions, free thiols present in proteins did not undergo reaction with pyrrole adducts to form cross-links. Further experiments using a low-molecular-mass pyrrole derivative indicated that glutathione may also able to suppress pyrrole dimerization without conjugate formation, possibly via inhibition of a free radical-dependent mechanism. The results suggest the following: (1) 2,5-HD-induced protein cross-linking is mediated primarily by pyrrole-to-pyrrole bridging under physiologic conditions, and (2) glutathione and other low-molecular-mass thiols may inhibit the pyrrole dimerization reaction by two distinct pathways. These findings have significant implications for the mechanism of gamma-diketone neuropathy.
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页码:764 / 771
页数:8
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