Bouchet M J, Jacques P, Ilien B, Goeldner M, Hirth C
Laborátoire de Chimie Bio-Organique, URA 1386 du CNRS, Faculté de Pharmacie, Université Louis Pasteur Strasbourg, Illkirch, France.
J Neurochem. 1992 Oct;59(4):1405-13. doi: 10.1111/j.1471-4159.1992.tb08454.x.
m-Sulfonate benzene diazonium chloride (MSBD) was used to affinity-label the gamma-aminobutyric acid (GABA) binding site from rat brain membranes. To assess the irreversibility of the labeling reaction, we used an efficient ligand dissociation procedure combined to a rapid [3H]muscimol binding assay, both steps being performed on filter-adsorbed membranes. Inactivation of specific [3H]-muscimol binding sites by MSBD and its prevention by GABA were both time- and concentration-dependent. The time course of MSBD labeling was shortened as the pH of the incubation medium was increased from 6.2 to 8. These data suggest that MSBD can efficiently label the GABA binding site through alkylation of a residue having an apparent dissociation constant around neutrality.
间磺酸苯重氮氯化物（MSBD）被用于亲和标记大鼠脑膜上的γ-氨基丁酸（GABA）结合位点。为评估标记反应的不可逆性，我们采用了一种高效的配体解离程序，并结合快速的[³H]蝇蕈醇结合测定法，这两个步骤均在滤膜吸附的膜上进行。MSBD对特异性[³H] - 蝇蕈醇结合位点的失活作用以及GABA对其的抑制作用均呈时间和浓度依赖性。随着孵育介质的pH从6.2提高到8，MSBD标记的时间进程缩短。这些数据表明，MSBD可以通过烷基化一个表观解离常数接近中性的残基来有效标记GABA结合位点。
