Waring A J, Chen Y, Faull K F, Stevens R, Sherman M A, Fluharty A L
Department of Pediatrics, Drew University-King Medical Center/University of California Los Angeles 90059, USA.
Mol Genet Metab. 1998 Jan;63(1):14-25. doi: 10.1006/mgme.1997.2646.
Cerebroside sulfate activator protein (CSAct or saposin B) is one of a group of heat stable, low-molecular-weight proteins that appear to share a common structural motif. These have been referred to as saposin-like proteins and are thought to share a multiple amphipathic helical barrel structure with a conserved pattern of disulfide linkages. Porcine kidney CSAct was prepared in high purity and consisted of three major glycosylated subforms. The protein was studied by physical-chemical methods and evaluated by various methods for structural prediction. All suggest that CSAct has high amounts of alpha-helical conformation and little if any beta-sheet. Circular dichroism (CD) studies indicate 45-50% helical conformation depending on buffer and temperature. There was only a moderate loss in helical content with increasing temperature and no indication of thermal denaturation. Fourier transform infrared spectroscopy (FTIR) measurements on deuterium hydrated self-films also indicated a predominantly helical structure. Helical axis orientation was investigated by both oriented CD and FTIR dichroism, which suggested that the helical axes were roughly parallel and oriented along the axis of the surface on which the self-films had been deposited. One-dimensional nuclear magnetic resonance spectra showed large chemical shift dispersion, indicating a defined tertiary structure with little variation between 6 and 85 degrees C. NOESY spectra failed to show the strong NOE cross peaks expected for a highly helical conformation. This may indicate short-term conformational flexibility within the helices or molecular aggregation at the high protein concentrations employed. These observations are consistent with the 3-4-helix bundle motif suggested for saposin-like proteins by various predictive algorithms.
硫酸脑苷脂激活蛋白（CSAct或鞘脂激活蛋白B）是一组热稳定的低分子量蛋白质之一，这些蛋白质似乎共享一个共同的结构基序。它们被称为类鞘脂激活蛋白，被认为共享一个具有保守二硫键连接模式的多重两亲性螺旋桶状结构。猪肾CSAct以高纯度制备，由三种主要的糖基化亚形式组成。通过物理化学方法对该蛋白质进行了研究，并通过各种结构预测方法进行了评估。所有结果都表明CSAct具有大量的α-螺旋构象，几乎没有β-折叠。圆二色性（CD）研究表明，根据缓冲液和温度的不同，螺旋构象为45%-50%。随着温度升高，螺旋含量仅有适度损失，且没有热变性的迹象。对氘水合自组装膜的傅里叶变换红外光谱（FTIR）测量也表明主要是螺旋结构。通过取向CD和FTIR二色性研究了螺旋轴取向，结果表明螺旋轴大致平行，并沿着自组装膜沉积的表面轴取向。一维核磁共振谱显示出较大的化学位移分散，表明在6至85摄氏度之间具有确定的三级结构且变化很小。NOESY谱未能显示出高度螺旋构象预期的强NOE交叉峰。这可能表明螺旋内存在短期构象灵活性，或者在所使用的高蛋白质浓度下存在分子聚集。这些观察结果与各种预测算法为类鞘脂激活蛋白提出的3-4螺旋束基序一致。
