The carbohydrate-binding domain on galectin-1 is more extensive for a complex glycan than for simple saccharides: implications for galectin-glycan interactions at the cell surface.
The carbohydrate-binding domain on galectin-1 is more extensive for a complex glycan than for simple saccharides: implications for galectin-glycan interactions at the cell surface.
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DOI：
10.1042/bj20090265
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发表时间：
2009-06-26
期刊：
影响因子：
--
通讯作者：
Mayo KH
中科院分区：
文献类型：
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作者：
Miller MC;Nesmelova IV;Platt D;Klyosov A;Mayo KH
Gal-1（半乳糖凝集素-1）主要通过与细胞表面糖缀合物中含有β-半乳糖苷的聚糖相互作用介导细胞-细胞和细胞-细胞外基质的粘附。尽管大多数关于gal-1的结构研究都研究了它与简单碳水化合物的结合，特别是乳糖和n -乙酰-乳胺，但这种观点是有限的，因为gal-1在细胞表面通过与更复杂的聚糖相互作用而起作用，这些聚糖在大小和组成上都是不均匀的。在本研究中，我们利用核磁共振光谱研究了人gal-1与一个大的（120 kDa）复合聚糖GRG （galactorhamnogalacturonate glycan）的相互作用，GRG含有非随机分布的大部分末端β（1→4）连接的半乳糖侧链。我们使用15N-1H-HSQC（异核单量子相干）核磁共振实验对富含15n的gal-1进行了鉴定，发现该区域覆盖了gal-1上很大的表面积，包括典型的乳糖结合位点，并从该位点穿过一个宽的谷或裂缝向二聚体界面延伸。HSQC和脉冲场梯度核磁共振扩散实验也表明，gal-1与GRG的化学计量比约为5:1（或6:1），平均宏观和微观平衡解离常数（Kd）分别为8×10−6 M和40×10−6 M（或48×10−6 M），比与乳糖的结合更强（Kd=520×10−6 M）。尽管gal-1可以以多种方式结合GRG，但乳糖可以竞争聚糖，这表明存在一种主要的相互作用模式。此外，尽管GRG上的末端基序是Gal-β(1→4)-Gal而不是传统的Gal-β(1→4)-Glc/GlcNAc（其中GlcNAc是n -乙酰氨基葡萄糖），但我们发现双糖Gal-β(1→4)-Gal可以在乳糖结合域与Gal- 1结合。此外，与GRG结合的gal-1破坏了聚糖间的相互作用，降低了聚糖介导的溶液粘度，这种聚糖去充血效应可能有助于解释为什么gal-1促进膜流动性和细胞膜内糖缀合物的横向扩散。总的来说，我们的研究结果提供了对原位凝集素功能的深入了解，并具有潜在的重大生物学后果。
gal-1 (galectin-1) mediates cell–cell and cell–extracellular matrix adhesion, essentially by interacting with β-galactoside-containing glycans of cell-surface glycoconjugates. Although most structural studies with gal-1 have investigated its binding to simple carbohydrates, in particular lactose and N-acetyl-lactosamine, this view is limited, because gal-1 functions at the cell surface by interacting with more complex glycans that are heterogeneous in size and composition. In the present study we used NMR spectroscopy to investigate the interaction of human gal-1 with a large (120 kDa) complex glycan, GRG (galactorhamnogalacturonate glycan), that contains non-randomly distributed mostly terminal β(1→4)-linked galactose side chains. We used 15N–1H-HSQC (heteronuclear single quantum coherence) NMR experiments with 15N-enriched gal-1 to identify the GRG-binding region on gal-1 and found that this region covers a large surface area on gal-1 that includes the quintessential lactose-binding site and runs from that site through a broad valley or cleft towards the dimer interface. HSQC and pulsed-field-gradient NMR diffusion experiments also show that gal-1 binds GRG with a gal-1:GRG stoichiometry of about 5:1 (or 6:1) and with average macroscopic and microscopic equilibrium dissociation constants (Kd) of 8×10−6 M and 40×10−6 M (or 48×10−6 M) respectively, indicating stronger binding than to lactose (Kd=520×10−6 M). Although gal-1 may bind GRG in various ways, the glycan can be competed for by lactose, suggesting that there is one major mode of interaction. Furthermore, even though terminal motifs on GRG are Gal-β(1→4)-Gal rather than the traditional Gal-β(1→4)-Glc/GlcNAc (where GlcNAc is N-acetylglucosamine), we show that the disaccharide Gal-β(1→4)-Gal can bind gal-1 at the lactose-binding domain. In addition, gal-1 binding to GRG disrupts inter-glycan interactions and decreases glycan-mediated solution viscosity, a glycan decongestion effect that may help explain why gal-1 promotes membrane fluidity and lateral diffusion of glycoconjugates within cell membranes. Overall, our results provide an insight into the function of galectin in situ and have potential significant biological consequences.