Fusion between-retinal rod outer segment membranes and model membranes: A role for photoreceptor peripherin/rds
Fusion between-retinal rod outer segment membranes and model membranes: A role for photoreceptor peripherin/rds
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DOI：
10.1021/bi980173p
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发表时间：
1998-06-30
期刊：
影响因子：
2.9
通讯作者：
Guo, YQ
中科院分区：
文献类型：
--
作者：
Boesze-Battaglia, K;Lamba, OP;Guo, YQ
Peripherin/rds在维持视杆细胞盘膜结构中起重要作用。将该蛋白质纯化至均一性[Boesze-Battaglia，K.，等人（1997）Biochemistry 36，6835-6846]使我们能够表征外周蛋白/rds在维持视杆外节（ROS）膜融合过程中的功能作用。利用无细胞融合检测系统，我们报告，融合的R-18-标记的ROS质膜（R-18-PM）与磁盘膜或外周/rds富集的大单层囊泡（LUVs）的外周/rds胰蛋白酶解抑制。为了理解这种现象，我们测试了一系列重叠的合成C-末端外周蛋白/rds肽介导模型膜融合的能力。在C-末端的63个氨基酸长的区域内，我们鉴定了促进膜融合所必需的最小15个残基长的氨基酸序列（PP-5）。PP-5能够抑制R-18-PM盘膜融合，并促进ANTS/DPX内容物在纯囊泡系统中的混合。这种肽（PP-5）促进钙诱导的囊泡聚集的磷脂酰乙醇胺：磷脂酰丝氨酸LUV。FTIR分析证实了该肽为α-螺旋的结构预测。当建模为α-螺旋时，该肽是两亲性的，疏水指数为0.75，疏水矩为0.59。PP-5与其他充分表征的膜融合蛋白具有实质性的生物化学和功能同源性。这些结果证明了外周蛋白/rds在ROS膜融合中的必要性，特别是该蛋白质的完整C-末端区域的需要。
Peripherin/rds plays an essential role in the maintenance of photoreceptor rod cell disk membrane structure. The purification of this protein to homogeneity [Boesze-Battaglia, K., et al. (1997) Biochemistry 36, 6835-6846] has allowed us to characterize the functional role of peripherin/rds in the maintenance of rod outer segment (ROS) membrane fusion processes. Utilizing a cell-free fusion assay system, we report that the fusion of R-18-labeled ROS plasma membrane (R-18-PM) with disk membranes or peripherin/rds-enriched large unilammellar vesicles (LUVs) is inhibited upon trypsinolysis of peripherin/rds. To understand this phenomenon, we tested the ability of a series of overlapping synthetic C-terminal peripherin/rds peptides to mediate model membrane fusion. Within the 63 amino acid long region of the C-terminus, we identified a minimal 15 residue long amino acid sequence (PP-5), which is necessary to promote membrane fusion. PP-5 was able to inhibit R-18-PM disk membrane fusion and promoted ANTS/DPX contents mixing in a pure vesicle system. This peptide (PP-5) promoted calcium-induced vesicle aggregation of phosphatidylethanolamine:phosphatidylserine LUVs. FTIR analysis confirmed the structural prediction of this peptide as alpha-helical. When modeled as an alpha-helix, this peptide is amphiphilic with a hydrophobicity index of 0.75 and a hydrophobic moment of 0.59. PP-5 has substantial biochemical and functional homology with other well-characterized membrane fusion proteins. These results demonstrate the necessity for peripherin/rds in ROS membrane fusion, specifically the requirement for an intact C-terminal region of this protein.