Natarajan K, Sawicki M W, Margulies D H, Mariuzza R A
Molecular Biology Section, Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Biochemistry. 2000 Dec 5;39(48):14779-86. doi: 10.1021/bi0018180.
CD69 is a widely expressed type II transmembrane glycoprotein related to the C-type animal lectins that exhibits regulated expression on a variety of cells of the hematopoietic lineage, including neutrophils, monocytes, T cells, B cells, natural killer (NK) cells, and platelets. Activation of T lymphocytes results in the induced expression of CD69 at the cell surface. In addition, cross-linking of CD69 by specific antibodies leads to the activation of cells bearing this receptor and to the induction of effector functions. However, the physiological ligand of CD69 is unknown. We report here the X-ray crystal structure of the extracellular C-type lectin-like domain (CTLD) of human CD69 at 2.27 A resolution. Recombinant CD69 was expressed in bacterial inclusion bodies and folded in vitro. The protein, which exists as a disulfide-linked homodimer on the cell surface, crystallizes as a symmetrical dimer, similar to those formed by the related NK cell receptors Ly49A and CD94. The structure reveals conservation of the C-type lectin-like fold, including preservation of the two alpha-helical regions found in Ly49A and mannose-binding protein (MBP). However, only one of the nine residues coordinated to Ca(2+) in MBP is conserved in CD69 and no bound Ca(2+) is evident in the crystal structure. Surprisingly, electron density suggestive of a puckered six-membered ring was discovered at a site structurally analogous to the ligand-binding sites of MBP and Ly49A. This sugar-like density may represent, or mimic, part of the natural ligand recognized by CD69.
CD69是一种广泛表达的II型跨膜糖蛋白，与C型动物凝集素相关，在造血谱系的多种细胞上呈现出受调控的表达，这些细胞包括中性粒细胞、单核细胞、T细胞、B细胞、自然杀伤（NK）细胞和血小板。T淋巴细胞的激活导致细胞表面CD69的诱导表达。此外，特定抗体对CD69的交联会导致携带该受体的细胞激活并诱导效应功能。然而，CD69的生理配体尚不清楚。我们在此报告人CD69细胞外C型凝集素样结构域（CTLD）在2.27埃分辨率下的X射线晶体结构。重组CD69在细菌包涵体中表达并在体外折叠。该蛋白在细胞表面以二硫键连接的同源二聚体形式存在，结晶为对称二聚体，类似于相关NK细胞受体Ly49A和CD94形成的二聚体。该结构揭示了C型凝集素样折叠的保守性，包括在Ly49A和甘露糖结合蛋白（MBP）中发现的两个α螺旋区域得以保留。然而，在MBP中与Ca(2+)配位的九个残基中只有一个在CD69中保守，并且在晶体结构中没有明显的结合Ca(2+)。令人惊讶的是，在一个结构上类似于MBP和Ly49A配体结合位点的位置发现了暗示有褶皱六元环的电子密度。这种类似糖的密度可能代表或模拟CD69识别的天然配体的一部分。
