Wray V, Federau T, Henklein P, Klabunde S, Kunert O, Schomburg D, Schubert U
Department of Molecular Structure Research, Institute of Biotechnological Research, Braunschweig, Germany.
Int J Pept Protein Res. 1995 Jan;45(1):35-43. doi: 10.1111/j.1399-3011.1995.tb01565.x.
The HIV-1 specific Vpu is a class I oligomeric membrane phosphoprotein of unknown structure and mechanism. The first experimental evidence for the position of secondary structural elements present in the hydrophilic C-terminal region of Vpu under various solution regimes is reported. CD data for nine overlapping 15 amino-acid fragments and 3 longer fragments indicate the presence of only transitory amounts of stable structure in aqueous solution alone, while with increasing trifluoroethanol content limiting structures were found indicating two helical segments in the hydrophilic region of Vpu. These limiting structures were more precisely defined from a detailed study of Vpu41-58, Vpu52-74 and Vpu63-81, by a combination of 2D 1H NMR spectroscopy, distance geometry, and restrained molecular dynamics and energy minimization calculations. Sets of low-energy conformations compatible with the quantitative NOE data indicate that Vpu41-58 has an alpha-helix from residues 42 to 50 while a second helix is found for Vpu52-74 from residues 57 to 69. Vpu63-81 shows only the presence of a single reverse turn at residues 74 to 77, without any evidence of helix, under the same conditions. From CD measurements the first helix extends back to residue 30 and is connected to the N-terminal anchor of Vpu. Thus the hydrophilic region of Vpu consists of two alpha-helices joined by a flexible region of 6 or 7 residues, which contains the phosphoacceptor sites of Vpu at positions 52 and 56. The second helix is followed by a single reverse turn and a flexible C-terminus.
HIV-1特异性Vpu是一种结构和作用机制未知的I类寡聚膜磷蛋白。本文报道了在不同溶液条件下，Vpu亲水C末端区域二级结构元件位置的首个实验证据。九个重叠的15个氨基酸片段和3个较长片段的圆二色性（CD）数据表明，仅在水溶液中仅存在少量短暂的稳定结构，而随着三氟乙醇含量的增加，发现了有限的结构，表明Vpu亲水区域存在两个螺旋段。通过二维1H核磁共振光谱、距离几何、受限分子动力学和能量最小化计算相结合的方法，对Vpu41-58、Vpu52-74和Vpu63-81进行了详细研究，更精确地定义了这些有限结构。与定量核Overhauser效应（NOE）数据兼容的低能量构象集表明，Vpu41-58在42至50位残基处有一个α螺旋，而Vpu52-74在57至69位残基处有第二个螺旋。在相同条件下，Vpu63-81仅在74至77位残基处存在一个单一的反向转角，没有任何螺旋的证据。通过CD测量，第一个螺旋延伸回30位残基，并与Vpu的N末端锚定相连。因此，Vpu的亲水区域由两个α螺旋组成，由6或7个残基的柔性区域连接，该柔性区域在52和56位含有Vpu的磷酸受体位点。第二个螺旋后面是一个单一的反向转角和一个柔性C末端。
