The cytoskeleton of a red blood cell (RBC) is anchored to the cell membrane by the ankyrin complex. This complex is assembled during RBC genesis and comprises primarily band 3, protein 4.2 and ankyrin, whose mutations contribute to numerous human inherited diseases. High-resolution structures of the ankyrin complex have been long sought-after to understand its assembly and disease-causing mutations. Here, we analyzed native complexes on the human RBC membrane by stepwise fractionation. Cryo-electron microscopy structures of nine band-3-associated complexes reveal that protein 4.2 stabilizes the cytoplasmic domain of band 3 dimer. In turn, the superhelix-shaped ankyrin binds to this protein 4.2 via ankyrin repeats (ARs) 6–13 and to another band 3 dimer via ARs 17–20, bridging two band 3 dimers in the ankyrin complex. Integration of these structures with both prior data and our biochemical data supports a model of ankyrin complex assembly during erythropoiesis and identifies interactions essential for the mechanical stability of RBC.

红细胞 (RBC) 的细胞骨架通过锚蛋白复合物锚定在细胞膜上。该复合物在红细胞形成过程中组装，主要包含带 3、蛋白 4.2 和锚蛋白，其突变导致许多人类遗传性疾病。锚蛋白复合物的高分辨率结构长期以来一直受到人们的追捧，以了解其组装和致病突变。在这里，我们通过逐步分级分离分析了人红细胞膜上的天然复合物。九个带 3 相关复合物的冷冻电镜结构表明，蛋白质 4.2 稳定带 3 二聚体的细胞质结构域。反过来，超螺旋形锚蛋白通过锚蛋白重复序列​​ (AR) 6-13 与该蛋白 4.2 结合，并通过 AR 17-20 与另一个带 3 二聚体结合，桥接锚蛋白复合物中的两个带 3 二聚体。