Boehm M K, Mayans M O, Thornton J D, Begent R H, Keep P A, Perkins S J
Department of Biochemistry and Molecular Biology, Royal Free Hospital of Medicine, London, UK.
J Mol Biol. 1996 Jun 21;259(4):718-36. doi: 10.1006/jmbi.1996.0353.
Carcinoembryonic antigen (CEA) is one of the most widely used cell-surface tumour markers for tumour monitoring and for targeting by antibodies. It is heavily glycosylated (50% carbohydrate) and a monomer is constructed from one V-type and six C2-type fold domains of the immunoglobulin superfamily. The solution arrangement at low resolution of the seven domains in CEA cleaved from its membrane anchor was determined by X-ray and neutron scattering. Guinier analyses showed that the X-ray radius of gyration RG of CEA was 8.0 nm. The length of CEA was 27 to 33 nm, and is consistent with an extended arrangement of seven domains. The X-ray cross-sectional radius of gyration RXS was 2.1 nm, and is consistent with extended carbohydrate structures in CEA. The neutron data gave CEA a relative molecular mass of 150,000, in agreement with a value of 152,500 from composition data, and validated the X-ray analyses. The CEA scattering curves were analysed using an automated computer modelling procedure based on the crystal structure of CD2. The V-type and C2-type domains in CD2 were separated, and the C2-type domain was duplicated five times to create a linear seven-domain starting model for CEA. A total of 28 complex-type oligosaccharide chains in extended conformations were added to this model. By fixing the six interdomain orientations to be the same, three-parameter searches of the rotational orientations between the seven domains gave 4056 possible CEA models. The best curve fits from these corresponded to a family of zig-zag models. The long axis of each domain was set at 160(+/-25) degrees relative to its neighbour, and the two perpendicular axes were orientated at 10(+/-30) degrees and -5(+/-35) degrees. Interestingly, the curve fit from this model is within error of that calculated from a CEA model generated directly from the CD2 crystal structure by the superposition of adjacent domains. Zig-zag models of this type imply that the protein face of the GFCC' beta-sheet in neighbouring CEA domains lie on alternate sides of the CEA structure. Such a model has implications for the adhesion interactions between CEA molecules on adjacent cells or for the antibody targeting of CEA.
癌胚抗原（CEA）是用于肿瘤监测和抗体靶向的应用最为广泛的细胞表面肿瘤标志物之一。它高度糖基化（50%为碳水化合物），一个单体由免疫球蛋白超家族的一个V型结构域和六个C2型折叠结构域构成。通过X射线和中子散射确定了从其膜锚定物上切割下来的CEA中七个结构域在低分辨率下的溶液排列。吉尼埃分析表明，CEA的X射线回转半径RG为8.0纳米。CEA的长度为27至33纳米，与七个结构域的伸展排列一致。X射线截面回转半径RXS为2.1纳米，与CEA中伸展的碳水化合物结构一致。中子数据得出CEA的相对分子质量为150,000，与组成数据得出的152,500的值相符，并验证了X射线分析结果。使用基于CD2晶体结构的自动计算机建模程序分析CEA散射曲线。将CD2中的V型和C2型结构域分离，并将C2型结构域复制五次，以创建CEA的线性七结构域起始模型。在此模型中总共添加了28条处于伸展构象的复合型寡糖链。通过固定六个结构域间的取向相同，对七个结构域之间的旋转取向进行三参数搜索，得到4056种可能的CEA模型。其中最佳的曲线拟合对应于一系列之字形模型。每个结构域的长轴相对于其相邻结构域设置为160（±25）度，两条垂直轴的取向分别为10（±30）度和 -5（±35）度。有趣的是，该模型的曲线拟合在误差范围内与通过相邻结构域叠加直接从CD2晶体结构生成的CEA模型计算得出的结果相符。这种之字形模型意味着相邻CEA结构域中GFCC'β - 折叠的蛋白质面位于CEA结构的交替两侧。这样的模型对相邻细胞上CEA分子之间的粘附相互作用或CEA的抗体靶向具有启示意义。
