Houen G, Koch C
Statens Seruminstitut, Department of Immunology, Copenhagen, S. Denmark.
Acta Chem Scand (Cph). 1994 Nov;48(11):905-11. doi: 10.3891/acta.chem.scand.48-0905.
Calreticulin is an intracellular protein known to be involved in calcium binding, but is also known to appear as an autoantigen in certain autoimmune diseases. The cDNA sequence is known but the protein has not yet been well characterized at the amino acid level. Owing to the possible involvement of this protein in autoimmune disease and with the aim of making monoclonal antibodies for use in assay development and immunohistochemistry, we have purified calreticulin using human placental material. Amino acid analysis of the purified protein confirmed the cDNA-derived composition, and only one discrepancy between the cDNA-predicted sequence and the amino acid sequence was found by peptide mapping and microsequencing. The protein contains one disulfide bridge and has one free SH group and the protein is neither glycosylated nor phosphorylated. Affinity chromatography of a placental protein extract on a column with immobilized calreticulin showed the existence of at least six proteins interacting with calreticulin. Using the purified calreticulin in Western blots, two out of eight patients with autoimmune disease diagnosed as having anti DNA antibodies in their serum were found also to contain autoantibodies to calreticulin in their serum.
钙网蛋白是一种细胞内蛋白质，已知其参与钙结合，但在某些自身免疫性疾病中也作为自身抗原出现。其cDNA序列已知，但该蛋白质在氨基酸水平上尚未得到充分表征。由于这种蛋白质可能与自身免疫性疾病有关，并且为了制备用于检测开发和免疫组织化学的单克隆抗体，我们使用人胎盘材料纯化了钙网蛋白。对纯化蛋白质的氨基酸分析证实了cDNA推导的组成，通过肽图谱和微量测序发现cDNA预测序列与氨基酸序列之间仅存在一个差异。该蛋白质含有一个二硫键，有一个游离的SH基团，且该蛋白质既不进行糖基化也不进行磷酸化。胎盘蛋白提取物在固定有钙网蛋白的柱上进行亲和层析，结果显示至少存在六种与钙网蛋白相互作用的蛋白质。在蛋白质印迹实验中使用纯化的钙网蛋白，发现八名血清中诊断为具有抗DNA抗体的自身免疫性疾病患者中有两名血清中也含有抗钙网蛋白自身抗体。
