BMP-3 and BMP-6 structures illuminate the nature of binding specificity with receptors.
Bone morphogenetic proteins (BMPs) are extracellular messenger ligands involved in controlling a wide array of developmental and intercellular signaling processes. To initiate their specific intracellular signaling pathways, the ligands recognize and bind two structurally related serine/threonine kinase receptors, termed type I and type II, on the cell surface. Here, we present the crystal structures of BMP-3 and BMP-6, of which BMP-3 has remained poorly understood with respect to its receptor identity, affinity, and specificity. Using surface plasmon resonance (BIAcore) we show that BMP-3 binds Activin Receptor type II (ActRII) with Kd approximately 1.8 microM but ActRIIb with 30-fold higher affinity at Kd approximately 53 nM. This low affinity for ActRII may involve Ser-28 and Asp-33 of BMP-3, which are found only in BMP-3's type II receptor-binding interfaces. Point mutations of either residue to alanine results in up to 20-fold higher affinity to either receptor. We further demonstrate by Smad-based whole cell luciferase assays that the increased affinity of BMP-3S28A to ActRII enables the ligand's signaling ability to a level comparable to that of BMP-6. Focusing on BMP-3's preference for ActRIIb, we find that Lys-76 of ActRII and the structurally equivalent Glu-76 of ActRIIb are distinct between the two receptors. We demonstrate that ActRIIbE76K and ActRII bind BMP-3 with similar affinity, indicating BMP-3 receptor specificity is controlled by the interaction of Lys-30 of BMP-3 with Glu-76 of ActRIIb. These studies illustrate how a single amino acid can regulate the specificity of ligand-receptor binding and potentially alter biological signaling and function in vivo.
Allendorph
George P
GP
Structural Biology Laboratory and Gene Expression Laboratory, Salk Institute for Biology Studies, 10010 North Torrey Pines Road, La Jolla, California 92037, USA.
Isaacs
Michael J
MJ
Kawakami
Yasuhiko
Y
Izpisua Belmonte
Juan Carlos
JC
Choe
Senyon
S
D055398:Bone Morphogenetic Protein 3; D055418:Bone Morphogenetic Protein 6; D019485:Bone Morphogenetic Proteins / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020349:Surface Plasmon Resonance
C525757:BMP3 protein, human; C525816:BMP6 protein, human; D055398:Bone Morphogenetic Protein 3; D055418:Bone Morphogenetic Protein 6; D019485:Bone Morphogenetic Proteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins