HIGH-AFFINITY PAROXETINE BINDING TO THE HUMAN PLACENTAL SEROTONIN TRANSPORTER
HIGH-AFFINITY PAROXETINE BINDING TO THE HUMAN PLACENTAL SEROTONIN TRANSPORTER
复制标题
DOI：
10.1152/ajpcell.1990.259.2.c196
复制
发表时间：
1990-08-01
影响因子：
--
通讯作者：
GANAPATHY, V
中科院分区：
文献类型：
--
作者：
COOL, DR;LEIBACH, FH;GANAPATHY, V
我们研究了非三环类抗抑郁药帕罗西汀与人胎盘刷状缘膜5-羟色胺转运体的相互作用。帕罗西汀以高亲和力结合在纯化的胎盘刷缘膜上[解离常数(Kd)=72 pM]。最大结合容量(Bmax)为3.9pmol/mg蛋白。丙咪嗪、地塞帕明和5-羟色胺以剂量依赖的方式抑制该结合，抑制常数值分别为4.4 nM、48.7 nM和1.77微米，而利血平、酮丝氨酸和5-羟色氨酸则无任何作用。丙咪嗪和5-羟色胺通过增加Kd值来抑制帕罗西汀的结合，但对Bmax基本没有影响。随着测定介质中Na+浓度的增加，帕罗西汀与膜的结合呈双曲线增加。CL-对结合力影响不大。Na+的作用主要是增加转运体对帕罗西汀的亲和力，而对Bmax无影响。缔合常数(Ka)随Na+浓度的增加呈双曲线增加，符合1Na+：1帕罗西汀的化学计量比。Ka的最大值为12.1+-。2.5次。1012M-1，Na+的Kd值为10.0+-。3.5毫米。用酪氨酸基特异性试剂处理膜减少了Na+依赖的结合，这表明酪氨酰残基参与了结合过程。然而，当试剂在Na+存在的情况下进行处理时，这种抑制作用显著降低，这表明活性酪氨酸残基位于或靠近Na+结合部位。在pH 6.5和7.5时，帕罗西汀均抑制胎盘刷状缘膜囊泡对氯化钠梯度依赖的5-羟色胺摄取。动力学分析表明，帕罗西汀增加转运体的表观解离常数(Kt)而不改变转运体的最大速度(Vmax)，因此在Nautre中这种抑制是竞争性的。结论：帕罗西汀是人胎盘5-羟色胺转运体的特异性高亲和力配体。
We investigated the interaction of paroxetine, a nontricyclic antidepressant, with the serotonin transporter of the human placental brush-border membrane. Paroxetine bound to the purified placental brush-border membranes with a high affinity [dissociation constant (Kd) = 72 pM]. The maximal binding capacity (Bmax) was 3.9 pmol/mg protein. Imipramine, desipramine, and serotonin inhibited the binding in a dose-dependent manner with inhibition constant (Ki) values of 4.4 nM, 48.7 nM, and 1.77 .mu.M, respectively, whereas reserpine, ketanserin, and 5-hydroxytryptophan did not have any effect. Imipramine and serotonin inhibite paroxetine binding by increasing the Kd with essentially no effect on Bmax. Binding of paroxetine to the membranes increased hyperbolically with increasing concentrations of Na+ in the assay medium. Cl- had little effect on the binding. The effect of Na+ was primarily to increase the affinity of the transporter for paroxetine with no effect on Bmax. The association constant (Ka) increased hyperbolically as the concentration of Na+ increased, indicating a 1Na+:1paroxetine stoichiometry. The maximal value for Ka was 12.1 .+-. 2.5 .times. 1012 M-1, and Kd for Na+ was 10.0 .+-. 3.5 mM. Treatment of the membranes with tyrosyl group-specific reagents reduced the Na+-dependent binding, suggesting the involvement of tyrosyl residues in the binding process. This inhibition was, however, significantly reduced when treatment with the reagent was performed in the presence of Na+, suggesting that the reactive tyrosyl residues were located at or near the Na+-binding site. Paroxetine inhibited NaCl gradient-dependent serotonin uptake in placental brush-border membrane vesicles both at pH 6.5 and 7.5. Kinetic analysis showed that the inhibition was competitive in nautre because paroxetine increased the apparent dissociation constant (Kt) of the transporter without altering the maximal velocity (Vmax). It is concluded that paroxetine is a specific high-affinity ligand for the human placental serotonin transporter.