ACETALDEHYDE ADDUCTS WITH PROTEINS - BINDING OF [C-14]-LABELED ACETALDEHYDE TO SERUM-ALBUMIN
ACETALDEHYDE ADDUCTS WITH PROTEINS - BINDING OF [C-14]-LABELED ACETALDEHYDE TO SERUM-ALBUMIN
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DOI：
10.1016/0003-9861(83)90406-x
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发表时间：
1983-01-01
影响因子：
3.9
通讯作者：
SORRELL, MF
中科院分区：
文献类型：
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作者：
DONOHUE, TM;TUMA, DJ;SORRELL, MF
乙醛是乙醇代谢的直接氧化产物，评估了其与溶液中纯化蛋白质共价结合的能力。牛血清白蛋白（BSA）用作模型蛋白，在 0.2 mM [ 14 C]乙醛存在下，pH 7.4，37℃下孵育。 C.乙醛与血清白蛋白形成稳定和不稳定的加合物。用还原剂硼氢化钠稳定后，鉴定出不稳定的加合物。使用三氯乙酸沉淀、凝胶过滤和透析作为分离结合乙醛和游离乙醛的方法来检查两种类型的结合。所有 3 种分析方法都给出了可比的结果，只是通过透析分离后，稳定的乙醛与白蛋白加合物的数量显着降低。评估了 L-半胱氨酸、L-赖氨酸和还原型谷胱甘肽作为竞争性试剂减少[14C]乙醛与 BSA 结合的能力。与赖氨酸引起的相比，半胱氨酸的添加导致[14C]乙醛与BSA的结合出现相当显着的浓度依赖性减少，而赖氨酸对共价结合表现出相对温和的影响。谷胱甘肽的存在导致蛋白质结合放射性的浓度依赖性降低，这种降低比赖氨酸更强，但不如半胱氨酸有效。还检查了每种试剂逆转与 BSA 预先形成的乙醛加合物形成的能力。只有 L-半胱氨酸有效地减少了乙醛与 BSA 的不稳定加合物的数量，而乙醛的稳定结合基本上不受 3 种试剂中任何一种的影响。乙醛可以与蛋白质共价结合并形成不稳定和稳定的加合物。
Acetaldehyde, the immediate oxidation production of ethanol metabolism, was assessed for its ability to bind covalently to a purified protein in solution. Bovine serum albumin (BSA) was used as the model protein incubated in the presence of 0.2 mM [14C]acetaldehyde at pH 7.4 nd at 37.degree. C. Acetaldehyde formed both stable and unstable adducts with serum albumin. Unstable adducts were identified following stabilization with the reducing agent sodium borohydride. Both types of binding were examined using trichloroacetic acid precipitation, gel filtration, and dialysis as means to separate bound from free acetaldehyde. All 3 methods of analysis gave comparable results except that the number of stable acetaldehyde adducts with albumin were significantly lower following separation by dialysis. The effects of L-cysteine, L-lysine, and reduced glutathione were assessed for their abilities as competitive reagents to decrease binding of [14C]acetaldehyde to BSA. Addition of cysteine caused a rather dramatic concentration-dependent reduction in [14C]acetaldehyde binding to BSA when compared to that caused by lysine which displayed a relatively mild effect on covalent binding. The presence of glutathione caused a concentration-dependent decrease in protein-bound radioactivity that was stronger than that by lysine but not as effective as cysteine. The ability of each reagent to reverse the formation of preformed acetaldehyde adducts with BSA was also examined. Only L-cysteine effectively decreased the number of unstable acetaldehyde adducts with BSA while stable binding of acetaldehyde remained essentially unaffected by any of the 3 reagents. Acetaldehyde can covalently bind to protein and form unstable as well as stable adducts.