Molecular recognition in purinergic receptors. 2. Diastereoselectivity of the h-P2Y1-receptor.
In the companion paper, part 1, we described the construction of an improved molecular model for the h-P2Y1 receptor (h-P2Y1-R) and proposed a rational for the stereoelectronic selectivity of the receptor. Here, we extend our studies on the molecular recognition of the h-P2Y1-R to the exploration of the diastereoselectivity of this receptor. For this purpose, we implemented an integrative approach combining synthesis, spectral analysis, biochemical assays, and computational analysis. Specifically, we selected and synthesized novel ATP analogues bearing a chiral center on the phosphate chain. We analyzed the conformation of the chiral ATP analogues in solution by 1H/13C NMR and assigned the absolute configuration of the diastereoisomers. The coordination mode of these analogues with a Mg2+ ion was evaluated by 31P NMR. These chiral analogues were biochemically evaluated and found to be potent h-P2Y1-R ligands. An EC50 difference of ca. 20-fold was observed between the diastereoisomers. Their spectral absolute configuration assignment was confirmed by comparison of the biochemical results to those of ATP-alpha-S diastereoisomers whose chirality is known. Finally, a computational analysis was performed for the elucidation of molecular recognition employing molecular mechanics (docking) studies on the receptor:ligands complexes. On the basis of the current results, we hypothesize that h-P2Y1-R's chiral discrimination originates from the requirement that the nucleotide analogue interacts with a Mg2+ ion within the receptor binding site. This Mg2+ ion is possibly coordinated with both Asp204 and the ATP's alpha, beta, gamma-phosphates in a Lambda configuration.
Major
Dan T
DT
Department of Chemistry, Gonda-Goldschmied Medical Research Center, Bar-Ilan University, Ramat-Gan 52900, Israel.
Nahum
Victoria
V
Wang
Yingfei
Y
Reiser
Georg
G
Fischer
Bilha
B
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000737:chemistry; D001665:Binding Sites; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D003198:Computer Simulation; D006801:Humans; D008024:Ligands; D008274:Magnesium / Q000737:chemistry; D008958:Models, Molecular; D008968:Molecular Conformation; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D018048:Receptors, Purinergic P2 / Q000737:chemistry* / Q000378:metabolism; D058472:Receptors, Purinergic P2Y1; D013237:Stereoisomerism; D013329:Structure-Activity Relationship
D008024:Ligands; C546382:P2RY1 protein, human; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D000255:Adenosine Triphosphate; D008274:Magnesium