Unique assignment of inter-subunit association in GABA(A) alpha1beta3gamma2 receptors determined by molecular modeling.

摘要

Recent publications defined requirements for inter-subunit contacts in a benzodiazepine-sensitive GABA(A) receptor (GABA(A)Ralpha1beta3gamma2). There is strong evidence that the heteropentameric receptor contains two alpha1, two beta3, and one gamma2 subunit. However, the available data do not distinguish two possibilities: When viewed clockwise from an extracellular viewpoint the subunits could be arranged in either gamma2beta3alpha1beta3alpha1 or gamma2alpha1beta3alpha1beta3 configurations. Here we use molecular modeling to thread the relevant GABA(A)R subunit sequences onto a template of homopentameric subunits in the crystal structure of the acetylcholine binding protein (AChBP). The GABA(A) sequences are known to have 15-18% identity with the acetylcholine binding protein and nearly all residues that are conserved within the nAChR family are present in AChBP. The correctly aligned GABA(A) sequences were threaded onto the AChBP template in the gamma2beta3alpha1beta3alpha1 or gamma2alpha1beta3alpha1beta3 arrangements. Only the gamma2alpha1beta3alpha1beta3 arrangement satisfied three known criteria: (1) alpha1 His(102) binds at the gamma2 subunit interface in proximity to gamma2 residues Thr(142), Phe(77), and Met(130); (2) alpha1 residues 80-100 bind near gamma2 residues 91-104; and (3) alpha1 residues 58-67 bind near the beta3 subunit interface. In addition to predicting the most likely inter-subunit arrangement, the model predicts which residues form the GABA and benzodiazepine binding sites.