A fluorescence study of type I and type II receptors of bone morphogenetic proteins with bis-ANS (4, 4'-dianilino-1, 1'-bisnaphthyl-5, 5' disulfonic acid).
Crystallography studies on several members of the bone morphogenetic protein (BMP) receptors suggested that hydrophobic regions in these proteins play an important role in their structure and function. In the present study, the environment sensitive fluorescent probe 4, 4'-dianilino-1, 1'-bisnaphthyl-5, 5' disulfonic acid (bis-ANS) was used to study the hydrophobic regions of the extracellular domain of the type I and II receptors for bone morphogenetic proteins (ecBMPR-IB and ecBMPR-II). A single bis-ANS binding site per receptor molecule was found for both receptors, but the two receptors interacted with bis-ANS with distinctive characteristics. A significant shift in the emission maximum from 498 to 510 nm was detected when bis-ANS binds ecBMPR-IB, but a negligible change in the emission maximum was observed when the dye binds ecBMPR-II. Under identical reaction conditions, the maximum fluorescence intensities of the probe (I(max)) for the ecBMPR-IB and -II are 4.0 and 6.2 x 10(4) arbitrary units, respectively. The probe binds to ecBMPR-IB and -II with K(d)=11.0 and 17.5 microM, respectively. The bis-ANS modified site on both receptor types was not readily accessible to acrylamide quenching. Fluorescence energy transfer experiments further revealed close proximity between the tyrosine (in ecBMPR-IB) and the tryptophan residue (in ecBMPR-II) and the respective bis-ANS binding site in these receptors. The binding of bis-ANS did not alter the ligand binding activity of ecBMPR-IB, but enhanced that of ecBMPR-II. These results show that the bis-ANS-modified hydrophobic site on the ecBMPR-IB and -II molecules plays a different functional role.
Yin
Huiran
H
Department of Biochemistry, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
Zhou
Qing
Q
Panda
Markandeswar
M
Yeh
Lee-Chuan C
LC
Zavala
Michelle C
MC
Lee
John C
JC
D000817:Anilino Naphthalenesulfonates / Q000737:chemistry*; D001665:Binding Sites; D052005:Bone Morphogenetic Protein Receptors, Type I / Q000737:chemistry*; D052006:Bone Morphogenetic Protein Receptors, Type II / Q000737:chemistry*; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D013050:Spectrometry, Fluorescence; D014443:Tyrosine / Q000737:chemistry
D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; D014443:Tyrosine; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); D052005:Bone Morphogenetic Protein Receptors, Type I; D052006:Bone Morphogenetic Protein Receptors, Type II