Interaction between soluble type I receptor for bone morphogenetic protein and bone morphogenetic protein-4.
Bone morphogenetic proteins (BMPs) are multifunctional proteins that comprise the largest subfamily of the transforming growth factor-beta. These proteins bind to types I and II serine/threonine kinase receptors. Ligand-induced heteromeric dimerization of these receptors is the key event in initiation of biological responses. We report here large-scale expression and purification of extracellular domain of the type I receptor for BMP-2/4, using a silkworm expression system. This soluble form of BMP receptor (sBMPR) was in monomer form in solution and bound to BMP-4 but not to activin or transforming growth factor-beta1. Surface plasmon resonance studies showed that kinetic parameters of sBMPR for BMP-4 consisted of a relatively rapid association rate constant (ka = 3.81 +/- 0.19 x 10(4) s-1 M-1) and an extremely slow dissociation rate constant (kd = 3.69 +/- 0.26 x 10(-4) s-1). From these two kinetic parameters, affinity was determined to be similar to that of the intact membrane-associated receptor expressed on COS cells. sBMPR inhibited the alkaline phosphatase activity in BMP responsive cell lines such as mouse osteoblastic cell MC3T3-E1 and bone marrow stromal cell ST2. These data indicate that the extracellular domain of type I receptor for BMP-2 and BMP-4 is sufficient for high-affinity binding to its ligands and should prove useful in understanding the role of BMP-2/4 in vivo, because a suitable high-affinity anti-BMP antibody has yet to be developed.
Natsume
T
T
Research and Development Center, Nippon Meat Packers, Inc., 3-3 Midorigahara, Tsukuba, Ibaraki 300-26, Japan. natsume@ims.u-tokyo.ac.jp
Tomita
S
S
Iemura
S
S
Kinto
N
N
Yamaguchi
A
A
Ueno
N
N
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D015374:Biosensing Techniques; D012831:Bombyx / Q000235:genetics; D001853:Bone Marrow / Q000378:metabolism; D001854:Bone Marrow Cells; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D052005:Bone Morphogenetic Protein Receptors, Type I; D019485:Bone Morphogenetic Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D017978:Receptors, Growth Factor / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012995:Solubility; D017154:Stromal Cells / Q000378:metabolism; D016212:Transforming Growth Factor beta
C525719:Bmp2 protein, mouse; C525810:Bmp4 protein, mouse; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D010446:Peptide Fragments; D017978:Receptors, Growth Factor; D011993:Recombinant Fusion Proteins; D016212:Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D052005:Bone Morphogenetic Protein Receptors, Type I