Identification of placenta growth factor determinants for binding and activation of Flt-1 receptor
Identification of placenta growth factor determinants for binding and activation of Flt-1 receptor
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DOI：
10.1074/jbc.m401418200
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发表时间：
2004-10-15
影响因子：
4.8
通讯作者：
De Falco, S
中科院分区：
文献类型：
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作者：
Errico, M;Riccioni, T;De Falco, S
胎盘生长因子(PlGF)属于血管内皮生长因子(VEGF)家族，是病理条件下血管生成事件的关键调节因子。PlGF通过结合和激活七个免疫球蛋白样域受体Flt-1，也称为VEGFR-1来发挥其生物学功能。在这里，我们报告了第一个详细的突变研究，为理解PlGF-1和Flt-1之间的分子识别提供了基础，重点介绍了一些对受体识别至关重要的残基。基于PlGF与Flt-1最小结合域相互作用的结构模型进行的突变分析，导致了与Flt-1识别有关的几个PlGF-1残基的鉴定。位于Beta3-Beta4环中的两个带负电荷的残基Asp-72和Glu-73对Flt-1的结合至关重要。其他导致PlGF结合活性显著下降的突变是位于N端α-螺旋的Gln-27，以及位于Beta6链上的Pro-98和Tyr-100。PlGF的两个糖基化残基之一ASN-84的突变产生了结合活性降低的PlGF变异体。这表明，与血管内皮生长因子不同，糖基化在Flt-1结合中起着重要作用。Asp-72和Glu-73残基的双重突变产生了一种PlGF变体，无法结合和激活细胞表面的受体分子。在体内的绒毛尿囊膜实验中，该变体未能在体外诱导原代内皮细胞的毛细血管样管形成或新生血管生成。
Placenta growth factor (PlGF) belongs to the vascular endothelial growth factor (VEGF) family and represents a key regulator of angiogenic events in pathological conditions. PlGF exerts its biological function through the binding and activation of the seven immunoglobulin-like domain receptor Flt-1, also known as VEGFR-1. Here, we report the first detailed mutagenesis studies that provide a basis for understanding molecular recognition between PlGF-1 and Flt-1, highlighting some of the residues that are critical for receptor recognition. Mutagenesis analysis, performed on the basis of a structural model of interaction between PlGF and the minimal binding domain of Flt-1, has led to the identification of several PlGF-1 residues involved in Flt-1 recognition. The two negatively charged residues, Asp-72 and Glu-73, located in the beta3-beta4 loop, are critical for Flt-1 binding. Other mutations, which bring about a significant decrease in PlGF binding activity, are Gln-27, located in the N-terminal alpha-helix, and Pro-98 and Tyr-100 on the beta6 strand. The mutation of one of the two glycosylated residues of PlGF, Asn-84, generates a PlGF variant with reduced binding activity. This indicates that, unlike in VEGF, glycosylation plays an important role in Flt-1 binding. The double mutation of residues Asp-72 and Glu-73 generates a PlGF variant unable to bind and activate the receptor molecules on the cell surface. This variant failed to induce in vitro capillary-like tube formation of primary endothelial cells or neo-angiogenesis in an in vivo chorioallantoic membrane assay.