HEXIX-TURN-HELIX PEPTIDES THAT FORM ALPHA-HELICAL FIBRILS

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 高密度脂蛋白大小的主要决定因素是其主要蛋白质载脂蛋白A-I（apo A-I）。 载脂蛋白A-I含有10个两亲性α-螺旋片段，这些片段依次相连。 我们合成了具有两个18-残基的两亲性α-螺旋片段的四种肽（设计为载脂蛋白A-I模型，Anantharamaiah，G. M.等）。由来自apo A-I的各种推定转角序列连接（1）。 1. Ac-DWLKAFYDKVAEKLKEAFKVEPLRADWLKAFYDKVAEKLKEAF-NH2 Ac-DWLKAFYDKVAEKLKEAFGLLPVLEDWLKAFYDKVAEKLKEAF-NH2 3. Ac-DWLKAFYDKVAEKLKEAFKVQPYLDDWLKAFYDKVAEKLKEAF-NH2 4. Ac-DWLKAFYDKVAEKLKEAFNGGARLADWLKAFYDKVAEKLKEAF-NH2 所有的肽形成不溶性聚集体后，在37？C在IOmM磷酸盐缓冲液（pH 7.6）中。 我们对这一观察结果很感兴趣，因为载脂蛋白A-I的变体与肝脏、心脏和肾脏的系统性淀粉样变性有关。 淀粉样变性是一种细胞外蛋白错误折叠起重要作用的疾病。 蛋白质聚集体作为b-折叠原纤维沉积在组织中。 3和4形成典型的淀粉样纤维，如硫磺素T和刚果红结合所示。 然而，与淀粉样蛋白原纤维相反，通过CD和FTIR，肽1和2形成α-螺旋原纤维，并且在溶液和原纤维膜中高度α-螺旋。 1和2不结合硫磺素T，并诱导刚果红光谱蓝移。 同步加速器X射线纤维衍射的磁性排列的样品1已证实的α-螺旋特性的原纤维。 衍射数据表明，与先前描述的卷曲螺旋原纤维（2-6）相反，α-螺旋垂直于主原纤维轴。据我们所知，由肽1形成的原纤维代表了一种新的结构。鉴于我们最近成功地与这些模型系统，肽1和其他（b-折叠形成）肽的进一步调查可能会显着提高我们的理解蛋白质折叠/错误折叠的性质。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. A major determinant of High Density Lipoprotein size is its main protein, Apolipoprotein A-I (apo A-I). Apo A-I contains 10 amphiphilic a-helical segments connected by turns. We synthesized four peptides with two 18-residue, amphiphilic a-helical segments (designed as apo A-I models, Anantharamaiah, G. M. et al.) connected by various putative turn sequences from apo A-I (1). 1. Ac-DWLKAFYDKVAEKLKEAFKVEPLRADWLKAFYDKVAEKLKEAF-NH2 2. Ac-DWLKAFYDKVAEKLKEAFGLLPVLEDWLKAFYDKVAEKLKEAF-NH2 3. Ac-DWLKAFYDKVAEKLKEAFKVQPYLDDWLKAFYDKVAEKLKEAF-NH2 4. Ac-DWLKAFYDKVAEKLKEAFNGGARLADWLKAFYDKVAEKLKEAF-NH2 All of the peptides formed insoluble aggregates after incubating at 37?C in 10 mM phosphate buffer, pH 7.6. We were intrigued by this observation because variants of apo A-I have been implicated in systemic amyloidosis of the liver, heart, and kidney. The amyloidoses consist of a large number of diseases in which misfolding of extracellular protein plays a prominent role. The protein aggregates are deposited in tissues as b-sheet fibrils. 3 and 4 formed typical amyloid fibrils, as shown by thioflavin T and Congo red binding. In contrast to amyloid fibrils, however, peptides 1 and 2 formed a-helical fibrils by CD and FTIR, and were highly a-helical in solution and in fibril films. 1 and 2 did not bind thioflavin T, and induced a blue shift in the spectrum of Congo red. Synchrotron x-ray fiber diffraction on a magnetically aligned sample of 1 has confirmed the a-helical character of the fibrils. The diffraction data suggest that the a-helices are perpendicular to the main fibril axis, in contrast to previously described coiled-coil fibrils (2-6). To the best of our knowledge, the fibrils formed from peptide 1 represent a novel structure. Given our recent success with these model systems, further investigations of peptide 1 and the other (b-sheet forming) peptides may significantly improve our understanding of protein folding / misfolding in nature.