Arya S, Chen F, Spycher S, Isenman D E, Shulman M J, Painter R H
Department of Biochemistry, University of Toronto, Ontario, Canada.
J Immunol. 1994 Feb 1;152(3):1206-12.
By analyzing the effects of single site mutations of a TNP-binding mouse IgM we have identified amino acid residues clustered in two regions in the C mu 3 domain that are important in the complement-dependent cytolytic activity of polymeric IgM. Some of the mutations also impaired IgM polymerization. For one of these clusters, D432G, P434A, and P436S, which lies on the fy2 and fy3 strands and their connecting loop, polymerization was little affected and the effect on the cytolytic activity of the polymer fraction was taken to imply direct involvement of the residue in C1 binding. The other cluster, involving residues D356A K361A and D417G, is situated at the other end of the C mu 3 domain closer to the center of the Fc mu disc. The D356A K361A and D417G mutations significantly impaired polymer formation, suggesting that these residues are necessary for proper folding or packing of the C mu 3 domains and may affect cytolysis only indirectly. Some other mutations had little or no effect on polymerization or cytolytic activity (E423A, E527G), whereas some mutations impaired only IgM polymerization without affecting cytolytic activity (D344A, K361A, K443A P544G). In others the defect in polymerization was so profound that only the monomer formed (H430A/N/Q and K438G). Our results also suggest that the C1 binding site of IgM is not strictly homologous to the C1 binding site of IgG. Although mutation of E318 of IgG has been shown to reduce its cytolytic activity, mutation of the homologous residue in IgM, E423, was without effect as were mutations of other flanking-charged residues. Proline at 436 in IgM and 331 in IgG may, however, be a common element.
通过分析一种结合TNP的小鼠IgM单一位点突变的影响，我们确定了Cμ3结构域中两个区域聚集的氨基酸残基，这些残基对聚合IgM的补体依赖性细胞溶解活性很重要。一些突变也损害了IgM聚合。对于位于fy2和fy3链及其连接环上的其中一个簇，即D432G、P434A和P436S，聚合受影响很小，并且对聚合物部分细胞溶解活性的影响被认为意味着该残基直接参与C1结合。另一个簇涉及残基D356A、K361A和D417G，位于Cμ3结构域的另一端，更靠近Fcμ盘的中心。D356A、K361A和D417G突变显著损害聚合物形成，表明这些残基对于Cμ3结构域的正确折叠或组装是必需的，并且可能仅间接影响细胞溶解。其他一些突变对聚合或细胞溶解活性几乎没有影响（E423A、E527G），而一些突变仅损害IgM聚合而不影响细胞溶解活性（D344A、K361A、K443A、P544G）。在其他情况下，聚合缺陷非常严重，以至于只形成了单体（H430A/N/Q和K438G）。我们的结果还表明，IgM的C1结合位点与IgG的C1结合位点并非严格同源。尽管已显示IgG的E318突变会降低其细胞溶解活性，但IgM中同源残基E423的突变没有影响，其他侧翼带电残基的突变也没有影响。然而，IgM中436位的脯氨酸和IgG中331位的脯氨酸可能是一个共同元素。
