Gadjeva Mihaela G, Rouseva Marieta M, Zlatarova Alexandra S, Reid Kenneth B M, Kishore Uday, Kojouharova Mihaela S
Channing Laboratory, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
Biochemistry. 2008 Dec 9;47(49):13093-102. doi: 10.1021/bi801131h.
The first step of activation of the classical complement pathway involves the binding of the globular C1q domain (gC1q) to the antigen-bound IgG or IgM. To improve our understanding of the mechanism of interaction of gC1q with IgG and IgM, we compared the immunoglobulin binding properties of single-residue mutants of individual globular modules of A and C chains. We found that Lys(A200) and Lys(C170) are significant for binding with both immunoglobulins. In addition, two C1q-specific scFv antibodies known as potent inhibitors of C1q-IgG and -IgM interactions were used in the epitope mapping analysis. A set of important residues, which participate in the C1q epitopes for scFv, were identified: Lys(C170) for the scFv3(V) epitope and Arg(B108) and Arg(B109) for the scFv10(V) epitope. The ability of scFv3(V) and scFv10(V) to bind preformed C1q-IgG or C1q-IgM complexes differed: scFv3(V) retained its ability to bind C1q, while scFv10(V) lost it. Given the different locations of the epitopes and the varying abilities of both antibodies to bind C1q-IgG and C1q-IgM complexes, we found that residues from the apical surface of C1q [where the scFv3(V) epitope was located] were involved in the initial recognition of IgG and IgM, while Arg(B108) and Arg(B109) are able to interact during the initial recognition as well as during the final binding of immunoglobulins. The reported results provide the first experimental evidence supporting the notion that apical and equatorial surfaces of gC1q have consecutive involvement following the gC1q reorientation during the interaction with specific C1q ligands.
经典补体途径激活的第一步涉及球形C1q结构域（gC1q）与抗原结合的IgG或IgM的结合。为了更好地理解gC1q与IgG和IgM相互作用的机制，我们比较了A链和C链单个球形模块的单残基突变体的免疫球蛋白结合特性。我们发现，赖氨酸（A200）和赖氨酸（C170）对于与两种免疫球蛋白的结合都很重要。此外，两种已知为C1q-IgG和-IgM相互作用有效抑制剂的C1q特异性单链抗体片段（scFv）被用于表位作图分析。鉴定出了一组参与scFv的C1q表位的重要残基：scFv3（V）表位的赖氨酸（C170）以及scFv10（V）表位的精氨酸（B108）和精氨酸（B109）。scFv3（V）和scFv10（V）结合预先形成的C1q-IgG或C1q-IgM复合物的能力有所不同：scFv3（V）保留了其结合C1q的能力，而scFv10（V）则失去了这种能力。鉴于表位的不同位置以及两种抗体结合C1q-IgG和C1q-IgM复合物的能力各异，我们发现C1q顶端表面（scFv3（V）表位所在位置）的残基参与了对IgG和IgM的初始识别，而精氨酸（B108）和精氨酸（B109）在免疫球蛋白的初始识别以及最终结合过程中都能够相互作用。所报道的结果提供了首个实验证据，支持了在与特定C1q配体相互作用期间gC1q重新定向后，gC1q的顶端和赤道表面连续参与的观点。
