Cheetham G M, Hale G, Waldmann H, Bloomer A C
MRC Laboratory of Molecular Biology, Hills Road, Cambridge, CB2 2QH, UK.
J Mol Biol. 1998 Nov 20;284(1):85-99. doi: 10.1006/jmbi.1998.2157.
The CAMPATH-1 family of antibodies are able systematically to lyse human lymphocytes with human complement by targeting the small cell-surface glycoprotein CD52, commonly called the CAMPATH-1 antigen. These antibodies have been used clinically for several years, providing therapy for patients with a variety of immunologically mediated diseases. We report here the first X-ray crystallographic analyses of a Fab fragment from a rat antibody, the original therapeutic monoclonal CAMPATH-1G and its humanized counterpart CAMPATH-1H, into which the six complementarity-determining regions of the rat antibody have been introduced. These structures have been refined at 2.6 A and 3.25 A resolution, respectively. The VL domains of adjacent molecules of CAMPATH-1H form a symmetric dimer within the crystals with an inter-molecular extended beta-sheet as seen in light chain dimers of the kappa class. Crystals of CAMPATH-1G have translational pseudo-symmetry. Within the antibody-combining sites, which are dominated by the protrusion of LysH52b and LysH53 from hypervariable loop H2, the charge distribution and overall integrity are highly conserved, but large changes in the position of loop H1 are observed and an altered conformation of loop H2. The major determinants of this are framework residues H71 and H24, whose identity differs in these two antibodies. These structures provide a detailed structural insight into the transplantation of an intact antibody-combining site between a rodent and a human framework, and provide an increased understanding of the specificity and antigen affinity of this pair of CAMPATH-1 antibodies for CD52. This study forms the structural basis for future modification and design of more effective antibodies to this important antigen.
CAMPATH-1家族抗体能够通过靶向小细胞表面糖蛋白CD52（通常称为CAMPATH-1抗原），利用人补体系统地裂解人淋巴细胞。这些抗体已在临床上使用多年，为患有多种免疫介导疾病的患者提供治疗。我们在此报告对大鼠抗体（原始治疗性单克隆抗体CAMPATH-1G）及其人源化对应物CAMPATH-1H的Fab片段进行的首次X射线晶体学分析，其中已引入了大鼠抗体的六个互补决定区。这些结构分别在2.6 Å和3.25 Å分辨率下进行了精修。CAMPATH-1H相邻分子的VL结构域在晶体内形成对称二聚体，具有分子间延伸的β折叠片层，如κ类轻链二聚体中所见。CAMPATH-1G的晶体具有平移假对称性。在抗体结合位点内，以高变环H2中LysH52b和LysH53的突出为主导，电荷分布和整体完整性高度保守，但观察到环H1位置有较大变化且环H2构象改变。主要决定因素是骨架残基H71和H24，它们在这两种抗体中的身份不同。这些结构为完整抗体结合位点在啮齿动物和人骨架之间的移植提供了详细的结构见解，并增进了对这对CAMPATH-1抗体对CD52的特异性和抗原亲和力的理解。这项研究为未来针对这一重要抗原设计更有效抗体的修饰和设计奠定了结构基础。
