Characterization of ligand-binding properties of the human BMP type II receptor extracellular domain.
ActR-IIA, ActR-IIB, and BMPR-II are low-affinity type II receptors that bind bone morphogenetic proteins (BMPs) in the same overall manner. The binding of BMPs by ActR-IIs has been analyzed structurally and functionally, but no detailed analysis of BMPR-II has been reported. The objective of this study was to determine ligand-binding epitopes and specificity determinants in two regions, the hydrophobic patch and the A-loop of the BMPR-II extracellular domain (ECD). A series of alanine-substituted variants was generated using a recently published X-ray structure of the unliganded form of the ovine BMPR-II ECD as a guide. These variants were characterized using one-dimensional NMR and functional activity assays with BMP-2, BMP-7 and GDF-5 as ligands. The results showed that alanine substitutions of conserved residues W85 and Y113 within the hydrophobic patch of the ECD differentially perturbed BMP ligand binding without disrupting receptor folding, suggesting that they are critical determinants for ligand binding and ligand specificity. Our results further revealed that the nonconserved residue L69 in the hydrophobic patch contributes to ligand-binding activity and specificity. Mutations of several residues within the A-loop resulted in minimal effects on the binding of the different BMP ligands. Overall, these observations identify several amino acid residues that play different roles in BMPR-II and ActR-II and thereby enable BMPR-II and ActR-IIs to bind different subclasses of BMP ligands.
Yin
Huiran
H
Department of Biochemistry, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX 78229-3900, USA.
Yeh
Lee-Chuan C
LC
Hinck
Andrew P
AP
Lee
John C
JC
D030301:Activin Receptors, Type II / Q000737:chemistry / Q000235:genetics; D000409:Alanine / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D052006:Bone Morphogenetic Protein Receptors, Type II / Q000737:chemistry* / Q000235:genetics; D017124:Conserved Sequence; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D017434:Protein Structure, Tertiary; D014364:Tryptophan / Q000737:chemistry / Q000235:genetics; D014443:Tyrosine / Q000737:chemistry / Q000235:genetics
D008024:Ligands; D014443:Tyrosine; D014364:Tryptophan; D030301:Activin Receptors, Type II; C498218:BMPR2 protein, human; D052006:Bone Morphogenetic Protein Receptors, Type II; D000409:Alanine