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将苹果 (MaluspumilaL .cv .Starkrimon)果肉微粒体和细胞可溶组分在含有3 H_ABA的缓冲介质中分别温育 ,仅在细胞可溶组分中测到微弱的3 H_ABA结合活性。但是 ,如果将果肉组织圆片在3 H_ABA缓冲介质中直接温育 ,经制备亚细胞组分后直接测定 ,在细胞可溶组分中测到很高的3 H_ABA特异结合活性。果肉圆片用沸水预先热处理使细胞可溶组分中的3 H_ABA结合活性完全丧失 ,说明ABA结合依赖于组织的活体状态。药理实验证明了ABA结合位点的蛋白质性质 ,同时证明该蛋白的活性中心具有—SH和丝氨酸基团。ABA结合蛋白对ABA的结合具有可饱和性、可逆性和高亲和力。Scatchard作图证明存在 2种ABA结合蛋白 ,一种具有较高的亲和力 ,其解离常数 (Kd)为 2 .9nmol/L ,另一种亲和力相对较低 ,其Kd值为 71.4nmol/L。用ABA结构相似物进行的竞争实验证明了ABA结合蛋白对配体结合的立体特异性。分析了ABA结合蛋白与ABA结合的时间曲线、pH和温度依赖性。本研究检测到的依赖活体组织的ABA结合蛋白可能是果实发育过程中介导ABA信号的受体。
Apple microspheres (Malus pumila L. cv. Starkrimon) and cell-soluble fractions were separately incubated in a buffer medium containing 3 H-ABA and only weak 3H-ABA-binding activity was detected in the cell-soluble fraction. However, if the pulp tissue discs were incubated directly in 3 H_ABA buffer medium and measured directly after production of the subcellular fractions, very high 3 HABA specific binding activity was detected in the cell-soluble fraction. Pre-heat treatment of the pulp slices with boiling water resulted in the complete loss of 3 H-ABA binding activity in the cell-soluble fraction, suggesting that ABA binding is dependent on the tissue state of the organism. Pharmacological experiments proved the protein nature of the ABA binding site, and at the same time proved that the active center of the protein has -SH and serine groups. The ABA binding protein has saturability, reversibility and high affinity for ABA binding. Scatchard mapping demonstrated the presence of two ABA binding proteins, one with high affinity with a dissociation constant (Kd) of 2.9 nmol / L and the other with a relatively low affinity with a Kd of 71.4 nmol / L. Competition experiments with ABA structural analogs demonstrate the stereospecificity of ABA binding proteins for ligand binding. The time course of ABA binding protein binding to ABA, pH and temperature dependence were analyzed. The ABA-binding proteins detected in this study may be the receptors that mediate ABA signaling during fruit development.