Molecular cloning of a regulatory protein for membrane-bound guanylate cyclase GC-A.
Molecular cloning of a regulatory protein for membrane-bound guanylate cyclase GC-A.
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膜结合鸟苷酸环化酶 GC-A 调节蛋白的分子克隆。
DOI：
10.1006/bbrc.2000.3761
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发表时间：
2000
影响因子：
3.1
通讯作者：
Chang,CH
中科院分区：
文献类型：
--
作者：
Chen,ZJ;Miao,ZH;Vetter,M;Dulin,N;Liu,S;Che,D;Hughes,B;Murad,F;Douglas,J;Chang,CH
心房利钠因子（ANF）激活膜结合鸟苷酸环化酶GC-A可能需要辅助蛋白的参与。为了确定这些假定的蛋白质，我们分离出一个1.0 kb的cDNA克隆从大鼠脑表达文库中使用的多克隆抗血清对mastoparan。该cDNA全长1.0 kb，编码111个氨基酸。该cDNA在COS-7细胞中的表达增强了ANF刺激的GC-A活性。因此，1.0 kb基因编码鸟苷酸环化酶调节蛋白（GCRP）。荧光显微镜下观察GCRP与绿色荧光蛋白（GFP）的融合蛋白表达，结果表明GCRP存在于PC 12细胞的胞浆中，但在ANF存在下，GCRP向质膜转移。免疫共沉淀实验表明，GCRP与GC-A在ANF的存在下。这些结果表明，ANF诱导GCRP与GC-A的缔合，并且这种缔合有助于GC-A的活化。
Activation of membrane-bound guanylate cyclase GC-A by atrial natriuretic factor (ANF) may require the involvement of accessory proteins. To identify these postulated proteins, we isolated a 1.0-kb cDNA clone from a rat brain expression library using a polyclonal antiserum against mastoparan. The 1.0-kb cDNA encodes a protein of 111 amino acids. Expression of this cDNA in COS-7 cells potentiated ANF-stimulated GC-A activity. Therefore, the 1.0-kb gene encodes a guanylate cyclase regulatory protein (GCRP). Fluorescence microscopy studies using the fusion protein of GCRP with green fluorescence protein (GFP) indicated that GCRP was present in the cytosol in PC12 cells, but translocated toward the plasma membrane in the presence of ANF. Coimmunoprecipitation experiments indicate that GCRP associates with GC-A in the presence of ANF. These results suggest that ANF induces the association of GCRP with GC-A and this association contributes to the activation of GC-A.