Molecular mechanism of cellular processing of transmembrane tumor necrosis factor

跨膜肿瘤坏死因子细胞加工的分子机制

• 批准号: 12660080
• 负责人: UTSUMI Toshihiko
• 金额: $ 2.3万
• 依托单位: Yamaguchi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12660080/
• 关键词: tumor necrosis factor (TNF); processing; posttranslational modification

Human transmembrane tumor necrosis factor (pro-TNF) was examined for posttranslational protein modification. The cDNA coding for pro-TNF was expressed in both COS-1 cells and Sf9 cells and metabolic labeling with [^3H]myristic or [^3H]palmitic acid was attempted. The 17 kDa mature TNF secreted from the transfected cells was not labeled, whereas the 26 kDa pro-TNF was specifically labeled with [^3H]palmitic acid. The [^3H]palmitic acid labeling of pro-TNF was eliminated by treatment with hydroxylamine, indicating that the labeling was due to palmitoylation of a cysteine residue via a thiocster bond. Site-directed mutagenesis of the two cysteine residues residing in the leader sequence of pro-TNF demonstrated that palmitoylation of pro-TNF occurs solely at Cys-47, located at the boundary between the transmembrane and cytoplasmic domains of pro-TNF.Thus, pro-TNF interacts with the plasma membrane via both its proteinaceous transmembrane domain and a lipid anchor.

检查人跨膜肿瘤坏死因子（Pro-TNF）是否进行翻译后蛋白质修饰。在COS-1细胞和SF9细胞中表达了编码Pro-TNF的cDNA，并尝试使用[^3H]肉豆蔻酸或[^3H]棕榈酸的代谢标记。未标记从转染细胞分泌的17 kDa成熟的TNF，而26 kDa pro-TNF则用[^3H]棕榈酸特异标记。通过用羟胺处理消除了pro-TNF的[^3H]棕榈酸标记，表明标记是由于通过硫代杆菌键对半胱氨酸残基的棕榈酰化引起的。位于Pro-TNF领导者序列中的两个半胱氨酸残基的位置定向诱变表明，pro-TNF的棕榈酰化仅发生在cys-47上，位于跨膜和细胞质域之间的边界上，pro-tnf的prostnf与prosmammammamma eNMBRANE的跨性别术语相互作用。

项目成果

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Utsumi, T.: "Transmembrane TNF (pro-TNF) is palmitoylated."FEBS Letters. 500. 1-6 (2001)

Utsumi, T.：“跨膜 TNF（TNF 前体）被棕榈酰化。”FEBS 快报。

Utsumi T, Takeshige T, Tanaka K, Takami K, Kira Y, Klostergaard J, Ishisaka R: "Transmembrane TNF (pro-TNF) is palmitoylated"FEBS Letters. 500. 1-6 (2001)

Utsumi T、Takeshige T、Tanaka K、Takami K、Kira Y、Klostergaard J、Ishisaka R：“跨膜 TNF（TNF 前体）被棕榈酰化”FEBS 快报。

Kashiwagi, A: "Suppression of T_3-and fatty acid-induced membrane permeability transition by L-carnitine"Comparative Biochemistry & Physiology. 130. 411-418 (2001)

柏木，A：“左旋肉碱抑制 T_3 和脂肪酸诱导的膜通透性转变”比较生物化学

Utsumi,T.: "Amino acid residue penultimate to amino-terminal Gly residue strongly affects two cotranslational protein modifications, N-myristoylation and N-acetylation."Journal of Biological Chemistry. (In press). (2001)

Utsumi,T.：“氨基末端甘氨酸残基倒数第二个氨基酸残基强烈影响两种共翻译蛋白修饰，N-肉豆蔻酰化和N-乙酰化。”《生物化学杂志》。