Attrill Helen, Takazawa Hirokazu, Witt Simone, Kelm Soerge, Isecke Rainer, Brossmer Reinhard, Ando Takayuki, Ishida Hideharu, Kiso Makoto, Crocker Paul R, van Aalten Daan M F
Division of Biological Chemistry, School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Biochem J. 2006 Jul 15;397(2):271-8. doi: 10.1042/BJ20060103.
Siglecs (sialic acid binding Ig-like lectins) are transmembrane receptors for sialylated glycoconjugates that modulate cellular interactions and signalling events in the haematopoietic, immune and nervous systems. Siglec-7 is a structural prototype for the recently described family of immune inhibitory CD33-related siglecs and is predominantly expressed on natural killer cells and monocytes, as well as subsets of CD8 T-cells. Siglec-specific inhibitors are desired for the detection of masked and unmasked forms of siglecs, to aid in dissection of signalling pathways and as tools to investigate siglecs as potential therapeutic targets. As a first step towards this end, we present the crystal structure of siglec-7 in complex with a sialylated ligand, the ganglioside analogue DSLc4 [alpha(2,3)/alpha(2,6) disialyl lactotetraosyl 2-(trimethylsilyl)ethyl], which allows for a detailed description of the binding site, required for structure-guided inhibitor design. Mutagenesis and binding assays were used to demonstrate a key structural role for Lys131, a residue that changes conformation upon sialic acid binding. Differences between the binding sites of siglec family members were then exploited using alpha-methyl Neu5Ac (N-acetylneuraminic acid) as a basic scaffold. A co-crystal of siglec-7 in complex with the sialoside inhibitor, oxamido-Neu5Ac [methyl alpha-9-(amino-oxalyl-amino)-9-deoxy-Neu5Ac] and inhibition data for the sialosides gives clear leads for future inhibitor design.
唾液酸结合免疫球蛋白样凝集素（Siglecs）是唾液酸化糖缀合物的跨膜受体，可调节造血、免疫和神经系统中的细胞相互作用和信号转导事件。Siglec-7是最近描述的免疫抑制性CD33相关Siglecs家族的结构原型，主要表达于自然杀伤细胞、单核细胞以及CD8 T细胞亚群上。检测Siglecs的隐蔽和非隐蔽形式、剖析信号通路以及作为研究Siglecs作为潜在治疗靶点的工具，都需要Siglec特异性抑制剂。作为实现这一目标的第一步，我们展示了Siglec-7与唾液酸化配体神经节苷脂类似物DSLc4[α(2,3)/α(2,6)二唾液酸乳糖四糖基2-(三甲基硅基)乙基]形成复合物的晶体结构，这使得能够详细描述结构导向抑制剂设计所需的结合位点。通过诱变和结合试验证明了Lys131的关键结构作用，该残基在唾液酸结合时会改变构象。然后以α-甲基Neu5Ac（N-乙酰神经氨酸）为基本支架，利用Siglec家族成员结合位点之间的差异。Siglec-7与唾液酸苷抑制剂草酰胺-Neu5Ac[甲基α-9-(氨基-草酰基-氨基)-9-脱氧-Neu5Ac]形成的共晶体以及唾液酸苷的抑制数据为未来的抑制剂设计提供了明确的线索。
