Transsulfuration Is a Significant Source of Sulfur for Glutathione Production in Human Mammary Epithelial Cells.
Transsulfuration Is a Significant Source of Sulfur for Glutathione Production in Human Mammary Epithelial Cells.
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DOI：
10.1155/2013/637897
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Wilkinson J 4th
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文献类型：
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作者：
Belalcázar AD;Ball JG;Frost LM;Valentovic MA;Wilkinson J 4th
来自甲硫氨酸循环的高半胱氨酸通过转硫途径为硫胱苷肽形成提供硫，硫胱苷肽随后可用于谷胱甘肽合成，迄今为止尚未鉴定为在乳腺细胞中有活性。原代人乳腺上皮细胞（HMEC）用S35-甲硫氨酸标记24小时，然后用载体对照、半胱氨酸生物合成抑制剂炔丙基甘氨酸或γ-谷氨酰半胱氨酸合成抑制剂丁硫氨酸亚砜亚胺预处理。制备细胞裂解物，并与谷胱甘肽-S-转移酶和荧光标记化合物一氯二亚胺反应以形成荧光谷胱甘肽-二亚胺缀合物。荧光和放射自显影图像的比较表明，谷胱甘肽已纳入S35-蛋氨酸证明功能转硫发生在乳腺细胞。途径抑制剂使掺入减少约80%。用和不用过氧化氢和/或途径抑制剂处理的HMEC中谷胱甘肽产生的测量表明，转硫途径在正常和氧化应激条件下为谷胱甘肽产生提供半胱氨酸方面起着重要作用。
The transsulfuration pathway, through which homocysteine from the methionine cycle provides sulfur for cystathionine formation, which may subsequently be used for glutathione synthesis, has not heretofore been identified as active in mammary cells. Primary human mammary epithelial cells (HMEC's) were labeled with S35-methionine for 24 hours following pretreatment with a vehicle control, the cysteine biosynthesis inhibitor propargylglycine or the gamma-glutamylcysteine synthesis inhibitor buthionine sulfoximine. Cell lysates were prepared and reacted with glutathione-S-transferase and the fluorescent labeling compound monochlorobimane to form a fluorescent glutathione-bimane conjugate. Comparison of fluorographic and autoradiographic images indicated that glutathione had incorporated S35-methionine demonstrating that functional transsulfuration occurs in mammary cells. Pathway inhibitors reduced incorporation by roughly 80%. Measurement of glutathione production in HMEC's treated with and without hydrogen peroxide and/or pathway inhibitors indicates that the transsulfuration pathway plays a significant role in providing cysteine for glutathione production both normally and under conditions of oxidant stress.