The crystal structure of placental growth factor in complex with domain 2 of vascular endothelial growth factor receptor-1.

Germaine Fuh,Hans W. Christinger,Christian Wiesmann,Abraham M. de Vos

doi:10.1074/jbc.m313237200

Abstract

Placental growth factor (PlGF) is a member of the vascular endothelial growth factor (VEGF) family and plays an important role in pathological angiogenic events. PlGF exerts its biological activities through binding to VEGFR1, a receptor tyrosine kinase that consists of seven immunoglobulin-like domains in its extracellular portion. Here we report the crystal structure of PlGF bound to the second immunoglobulin-like domain of VEGFR1 at 2.5 A resolution and compare the complex to the closely related structure of VEGF bound to the same receptor domain. The two growth factors, PlGF and VEGF, share a sequence identity of approximately 50%. Despite this moderate sequence conservation, they bind to the same binding interface of VEGFR1 in a very similar fashion, suggesting that both growth factors could induce very similar if not identical signaling events.

Highlights

The atomic coordinates and structure factors have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ
Placental growth factor (PlGF) exerts its biological activities through binding to VEGFR1, a receptor tyrosine kinase that consists of seven immunoglobulin-like domains in its extracellular portion
We report the crystal structure of PlGF bound to the second immunoglobulin-like domain of VEGFR1 at 2.5 Å resolution and compare the complex to the closely related structure of vascular endothelial growth factor (VEGF) bound to the same receptor domain

Summary

EXPERIMENTAL PROCEDURES

Expression, Refolding, and Purification—VEGFR constructs were expressed, purified, and tested for functional integrity as described [17, 19]. The concentrations of a VEGFR1 fragment that produced a signal on a VEGF-conjugated chip over a blank chip in the range of 30 to 70 response units in a 2-min injection at 30 ␮l/min were determined. The amount of captured RU was plotted against the concentrations of VEGF or PlGF in the initial incubation stage, and IC50 levels were determined This assay was restricted by the lowest concentrations of VEGFR1 fragments sufficient to produce a reasonable signal. In these instances we report the affinity derived from other assays such as radioimmunoreceptor competition binding assays or biotinylated protein enzymelinked immunosorbent assays with a more sensitive readout as described previously [17]. Using data from 12 to 6 Å yielded a clear

TABLE I Binding affinities

RESULTS

TABLE II Data collection and refinement statistics

DISCUSSION