Brain G protein gamma subunits contain an all-trans-geranylgeranylcysteine methyl ester at their carboxyl termini.
Brain G protein gamma subunits contain an all-trans-geranylgeranylcysteine methyl ester at their carboxyl termini.
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脑G蛋白γ亚基在其羧基末端含有全反式香叶基香叶基半胱氨酸甲酯。
DOI：
10.1073/pnas.87.15.5868
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发表时间：
1990
影响因子：
11.1
通讯作者：
Glomset,JA
中科院分区：
文献类型：
--
作者：
Yamane,HK;Farnsworth,CC;Xie,HY;Howald,W;Fung,BK;Clarke,S;Gelb,MH;Glomset,JA
我们先前已经表明，从牛脑膜纯化的鸟嘌呤核苷酸结合蛋白（G蛋白）β-γ复合物在γ多肽的C-末端半胱氨酸残基上被甲酯化。在本研究中，3 H-甲基化的G β γ复合物裂解其组成氨基酸的穷举蛋白水解被证明含有放射性标记的材料，共洗脱与香叶基香叶基半胱氨酸甲酯的反相HPLC和两个TLC系统。通过过甲酸氧化的进一步处理产生与L-半胱氨酸甲酯共洗脱的放射性标记材料，验证异戊二烯基修饰发生在C-末端半胱氨酸残基上。通过气相色谱-质谱联用分析，从纯化的G β γ通过处理与雷尼镍释放的材料，积极确定了共价结合的脂质作为一个全反式-香叶基香叶基（C20）类异戊二烯部分。为了描述这种修饰在γ亚基中的分布，通过反相HPLC将纯化的G β γ复合物分离成5-kDa（γ 5）和6-kDa（γ 6）形式的γ多肽。气相色谱-质谱联用分析的雷尼镍处理的纯化γ 5和γ 6亚基表明，这两种多肽被修改的geranylgeranylation。这些结果表明，至少有两种形式的脑γ亚基的后修饰的牛儿基牛儿基化和羧基甲基化。这些修饰对于将G β γ复合物靶向膜可能是重要的。
We have shown previously that guanine nucleotide-binding protein (G protein) beta gamma complexes purified from bovine brain membranes are methyl esterified on a C-terminal cysteine residue of the gamma polypeptide. In the present study, 3H-methylated G beta gamma complexes cleaved to their constituent amino acids by exhaustive proteolysis were shown to contain radiolabeled material that coeluted with geranylgeranylcysteine methyl ester on reversed-phase HPLC and two TLC systems. Further treatment by performic acid oxidation yielded radiolabeled material that coeluted with L-cysteic acid methyl ester, verifying that the prenyl modification occurs on a C-terminal cysteine residue. Analysis by gas chromatography-coupled mass spectrometry of material released from purified G beta gamma by treatment with Raney nickel positively identified the covalently bound lipid as an all-trans-geranylgeranyl (C20) isoprenoid moiety. To delineate the distribution of this modification among gamma subunits, purified G beta gamma complexes were separated into 5-kDa (gamma 5) and 6-kDa (gamma 6) forms of the gamma polypeptide by reversed-phase HPLC. Gas chromatography-coupled mass spectrometry analyses of Raney nickel-treated purified gamma 5 and gamma 6 subunits showed that both polypeptides were modified by geranylgeranylation. These results demonstrate that at least two forms of brain gamma subunit are posttranslationally modified by geranylgeranylation and carboxyl methylation. These modifications may be important for targeting G beta gamma complexes to membranes.