COVALENT MODIFICATION OF LYSINE DURING THE SUICIDE INACTIVATION OF RAT-LIVER CYTOCHROME-P-450 BY CHLORAMPHENICOL
COVALENT MODIFICATION OF LYSINE DURING THE SUICIDE INACTIVATION OF RAT-LIVER CYTOCHROME-P-450 BY CHLORAMPHENICOL
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DOI：
10.1016/s0006-2952(81)80010-x
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发表时间：
1981-01-01
影响因子：
5.8
通讯作者：
HALPERT, J
中科院分区：
文献类型：
--
作者：
HALPERT, J
在重组的单加氧酶系统或苯巴比妥处理的大鼠完整肝微粒体代谢抗生素[14C]氯霉素的过程中，含有14C的代谢物共价结合到细胞色素P-450上，该酶被不可逆地抑制。在这两个系统中，.apprx。与蛋白质共价结合的14C中，95%与细胞色素P-450有关。在0.5%十二烷基硫酸钠存在下，用1N羟胺(pH 7.5)或温和碱性水解液(pH 10.5)解离与重组体系蛋白质结合的~(14)C的一半。在用蛋白水解酶消化P-450的条件下，大多数羟胺不稳定的蛋白结合物质很容易被降解成草酸。从重组体系中~(14)C标记的蛋白质或~(14)C标记的微体蛋白的链霉蛋白酶消化液中分离到对羟胺稳定的放射性标记物质作为氨基酸加合物。根据与合成化合物的共层析和从微粒体蛋白的消化液中分离的14C产物碱性水解时游离赖氨酸和氯霉素草酸的释放，该加合物被鉴定为N-β-氯霉素草酰赖氨酸。赖氨酸的修饰是氯霉素对细胞色素P-450自杀性失活的部分原因。
During the metabolism of the antibiotic [14C]chloramphenicol by a reconstituted monooxygenase system or by intact liver microsomes from phenobarbital-treated rats, a 14C-containing metabolite covalently bonded to cytochrome P-450 and the enzyme was irreversibly inhibited. In both systems, .apprx. 95% of the 14C that was covalently bound to protein was associated with cytochrome P-450. In the presence of 0.5% sodium dodecylsulfate, half of the 14C that was bound to the proteins of the reconstituted system was dissociated by treatment with 1 N hydroxylamine (pH 7.5) or by mild alkaline hydrolysis (pH 10.5). Most of the hydroxylamine-labile protein-bound material was readily degraded to oxalate under the conditions used to digest the P-450 with proteolytic enzymes. The radiolabeled material stable to hydroxylamine was isolated as an amino acid adduct from pronase digests of the 14C-labeled proteins of a reconstituted system or the 14C-labeled microsomal proteins. This adduct was identified as N-.epsilon.-chloramphenicol oxamyl lysine based on co-chromatography with the synthetic compound and on the release of free lysine plus chloramphenicol oxamic acid upon alkaline hydrolysis of the 14C product isolated from digests of the microsomal protein. Modification of lysine is partially responsible for the suicide inactivation of cytochrome P-450 by chloramphenicol.