DOUBLY-LIPID-MODIFIED PROTEIN-SEQUENCE MOTIFS EXHIBIT LONG-LIVED ANCHORAGE TO LIPID BILAYER-MEMBRANES
DOUBLY-LIPID-MODIFIED PROTEIN-SEQUENCE MOTIFS EXHIBIT LONG-LIVED ANCHORAGE TO LIPID BILAYER-MEMBRANES
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DOI：
10.1021/bi00011a039
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发表时间：
1995-03-21
期刊：
影响因子：
2.9
通讯作者：
SILVIUS, JR
中科院分区：
文献类型：
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作者：
SHAHINIAN, S;SILVIUS, JR
为了更好地理解在真核生物的一些细胞内蛋白质中发现的双脂质修饰的潜在功能重要性，我们已经研究了如何“顽强地”各种双脂质修饰的肽，其序列和脂质修饰反映了细胞内蛋白质中发现的那些，被锚定到脂质双层膜上。将带有双脂质修饰的荧光标记肽掺入大的单层卵磷脂/磷脂酰甘油囊泡中，并通过荧光去猝灭试验监测脂肽的自发囊泡间转移的动力学。掺入稳定的“双锚”基序的脂肽-C（香叶基香叶基）XC在几种rab和同源蛋白中发现的（香叶基香叶基）-OMe表现出非常慢的双层间转移速率（t（1/2）50 h），与掺入肉豆蔻酰-GC的脂肽一样（棕榈酰）X-和-C（棕榈酰）XC（法呢基）-OMe基序分别存在于多种src相关的细胞内酪氨酸激酶和G蛋白α亚基以及p21（H-ras）中。终止于甲基化-C（香叶基香叶基）-OH基序的脂肽显示出稍微更大但仍然非常慢的自发双层间转移速率（t（1/2）= ca. 10 h）。从这些结果推断，我们估计相应的双锚定蛋白从膜上自发解吸的速率应该比受调节的蛋白介导的释放速率慢得多（通过与“护送"蛋白结合或通过去S-酰化来实现），因此，这些物种的细胞内分布细胞内的蛋白质（特别是它们靶向特定的细胞内膜）可能主要由动力学因素而不是热力学因素支配（和调节）。在平行的囊泡结合测量，与S-棕榈酰基团修饰的肽被发现与脂质双层相关联，甚至更贪婪地比可比geranylgeranylated肽，解释了“顽强的”膜结合特性的双锚基序纳入一个S-棕榈酰残基。
To understand better the potential functional importance of the dual-lipid modifications found in a number of intracellular proteins of eukaryotes, we have examined how ''tenaciously'' various doubly-lipid-modified peptides, with sequences and lipid modifications reflecting those found in intracellular proteins, are anchored to lipid bilayer membranes. Fluorescent-labeled peptides bearing dual-lipid modifications were incorporated into large unilamellar egg phosphatidylcholine/phosphatidylglycerol vesicles, and the kinetics of spontaneous intervesicle transfer of the lipopeptides were monitored by a fluorescence-dequenching assay. Lipopeptides incorporating the stable ''dual-anchor'' motif -C(geranylgeranyl)XC(geranylgeranyl)-OMe found in several rab and homologous proteins exhibit very slow rates of interbilayer transfer (t(1/2) 50 h), as do lipopeptides incorporating myristoyl-GC(palmitoyl)X- and -C(palmitoyl)XC(farnesyl)-OMe motifs found in various src-related intracellular tyrosine kinases and G-protein alpha-subunits and in p21(H-ras), respectively. Lipopeptides terminating in an ummethylated -C(geranylgeranyl)C(geranylgeranyl)-OH motif show somewhat greater but still very slow rates of spontaneous interbilayer transfer (t(1/2) = ca. 10 h). Extrapolating from these results, we estimate that the rate of spontaneous desorption of the corresponding doubly-anchored proteins from membranes should be much slower than that Of regulated, protein-mediated release (effected by binding to an ''escort'' protein Or by de-S-acylation), As a result the intracellular distributions of these species (and particularly their targeting to specific intracellular membranes) are likely to be governed (and regulated) primarily by kinetic rather than thermodynamic factors. In parallel vesicle-binding measurements, peptides modified with S-palmitoyl groups were found to associate with lipid bilayers even more avidly than comparable geranylgeranylated peptides, explaining the 'tenacious' membrane-binding properties of dual-anchor motifs incorporating an S-palmitoyl residue.