Structural and functional basis of CXCL12 (strornal cell-derived factor-1α) binding to heparin
Structural and functional basis of CXCL12 (strornal cell-derived factor-1α) binding to heparin
复制标题
DOI：
10.1074/jbc.m608796200
复制
发表时间：
2007-03-30
影响因子：
4.8
通讯作者：
Lolis, Elias
中科院分区：
文献类型：
--
作者：
Murphy, James W.;Cho, Yoonsang;Lolis, Elias
CXCL12(SDF-1α)和CXCR4在成人胚胎发育和细胞迁移中起关键作用。这些蛋白质与HIV-1感染、癌症转移和突发疾病有关。糖胺多聚糖(GAG)对CXCR4上CXCL12的截留和递呈对受体的激活具有重要作用。突变技术已经确定了CXCL12残基与肝素结合。然而，这种相互作用的分子细节还没有确定。在这里，我们证明了可溶性肝素和硫酸肝素对CXCL12介导的体外趋化作用有负面影响。我们还表明，二聚体界面上的一簇碱性残基是趋化所必需的，也是肝素抑制的目标。我们提出了不饱和肝素二糖与CXCL12结合的结构证据，这是通过溶液核磁共振光谱和X射线结晶学获得的。随着二糖浓度的增加，CXCL12的二维H-1-N-15-HSQC谱发生了变化，确定了两个残基簇。一个簇对应于二聚体界面中的β-链。第二个包括氨基末端环和α-螺旋。在x射线结构中，存在两种不饱和双糖。一种是CXCL12的His(25)Lys(27)和Arg(41)残基与肝素二糖直接接触的二聚体界面。第二个二糖与丙氨酸(20)、精氨酸(21)、天冬氨酸(30)和赖氨酸()接触。这是第一个CXC类趋化因子与糖胺聚糖形成的复合体的X射线结构。基于对两个肝素结合位点的观察，我们提出了一种机制，即GAG在CXCL12二聚体隔离并将CXCL12呈现给CXCR4时与其结合。
CXCL12 (SDF-1 alpha) and CXCR4 are critical for embryonic development and cellular migration in adults. These proteins are involved in HIV-1 infection, cancer metastasis, and WHIM disease. Sequestration and presentation of CXCL12 to CXCR4 by glycosaminoglycans (GAGs) is proposed to be important for receptor activation. Mutagenesis has identified CXCL12 residues that bind to heparin. However, the molecular details of this interaction have not yet been determined. Here we demonstrate that soluble heparin and heparan sulfate negatively affect CXCL12-mediated in vitro chernotaxis. We also show that a cluster of basic residues in the dimer interface is required for chemotaxis and is a target for inhibition by heparin. We present structural evidence for binding of an unsaturated heparin disaccharide to CXCL12 attained through solution NMR spectroscopy and x-ray crystallography. Increasing concentrations of the disaccharide altered the two-dimensional H-1-N-15-HSQC spectra of CXCL12, which identified two clusters of residues. One cluster corresponds to beta-strands in the dimer interface. The second includes the amino-terminal loop and the a-helix. In the x-ray structure two unsaturated disaccharides are present. One is in the dimer interface with direct contacts between residues His(25) Lys(27), and Arg(41) of CXCL12 and the heparin disaccharide. The second disaccharide contacts Ala(20), Arg(21), Asn(30), and Lys(64). This is the first x-ray structure of a CXC class chemokine in complex with glycosaminoglycans. Based on the observation of two heparin binding sites, we propose a mechanism in which GAGs bind around CXCL12 dimers as they sequester and present CXCL12 to CXCR4.