Monospecific antibodies as probes for the stoichiometry of recombinant GABA(A) receptors.

Knight, A.R., Stephenson, F.A., Tallman, J.F. and Ramabahdran, T.V. (2000) Monospecific antibodies as probes for the stoichiometry of recombinant GABA(A) receptors. Receptors & Channels, 7 (3). pp. 213-226.

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Abstract

GABA(A) receptors composed of alpha1beta3 gamma2 and alpha1beta3 subunits were expressed in insect Sf9 cells and solubilized in 1% Triton X100. In sucrose density gradients, [3H]-Ro15-1788 binding activity, in the case of alpha1beta3 gamma2, and [3H]-muscimol binding activity, in the case of alpha1beta3 containing receptors sedimented as a single sharp peak suggesting the formation of receptors containing a defined number of subunits. When alpha1beta3gamma2 -containing receptors were incubated with an alpha-subunit specific antibody (bd24), a single class of antibody receptor complex was formed irrespective of the receptor-antibody ratio. This is consistent with two alpha subunits cross-linked within the receptor by the antibody. Similar results were obtained using a beta-subunit specific antibody (bd17). Several classes of antibody-receptor complex were formed when receptors were pre-incubated with a gamma specific antibody (anti gamma(2) 1-15 Cys). This profile is consistent with the presence of a single gamma subunit in each complex. Experiments with alpha1beta3 subunit containing receptors and antibody bd24 produced a profile similar to that seen with alpha1beta3 gamma2 receptors, consistent with two alpha subunits per receptor complex. In this case, the anti-beta subunit antibody, bd17, produced a unique and complex profile consistent with three beta subunits per receptor. This method permits the rapid determination of subunit stoichiometries of homogeneous receptor populations

Item Type:Article
Departments, units and centres:Department of Pharmaceutical and Biological Chemistry > Department of Pharmaceutical and Biological Chemistry
ID Code:2540
Journal or Publication Title:Receptors & Channels
Deposited By:Library Staff
Deposited On:25 Nov 2011 10:52
Last Modified:25 Nov 2011 10:52

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