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首页> 外文期刊>Biochemistry >Domain-specific stabilization of photoreceptor membrane guanylyl cyclase by adenine nucleotides and guanylyl cyclase activating proteins (GCAPs)
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Domain-specific stabilization of photoreceptor membrane guanylyl cyclase by adenine nucleotides and guanylyl cyclase activating proteins (GCAPs)

机译：腺嘌呤核苷酸和鸟苷酸环化酶激活蛋白（GCAP）稳定感光膜鸟苷酸环化酶的域特异性

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摘要

In photoreceptor outer segments, particulate guanylyl cyclase (RetGC) is stimulated at low intracellular Ca2+ concentrations by guanylyl cyclase activating protein (GCAP), a Ca2+-sensitive activator, to resynthesize light-depleted cGMP. In washed outer segment membranes, we find that GCAP-stimulable RetGC is rapidly inactivated at physiological temperatures (30-37 degrees C). Under the same conditions, RetGC remains competent for stimulation by S-100 protein preparations or Mn2+/Triton X-100, indicating that the cyclase catalytic domain remains functional. GCAPs and adenine nucleotides protect against inactivation. Protection by GCAPs is independent of Ca2+ concentration, suggesting that there is a Ca2+-independent interaction between GCAP and RetGC. Protection by ATP (EC50 = 150 microM) is not due to phosphorylation, since the nonhydrolyzable analogue adenylyl imidodiphosphate (AMP-PNP) protects equally well. In addition to their roles in protection, ATP and AMP-PNP also slowly stimulate cyclase activity. In parallel with the functional change in RetGC at physiological temperatures, we also observe a structural change. A 62-kDa intracellular fragment of RetGC-1 becomes more sensitive to cleavage by trypsin after preincubation at 30 degrees C unless ATP, AMP-PNP, or GCAP is present. Adenine nucleotides and GCAPs thus protect RetGC structurally, as well as functionally.
机译：在感光器外部片段中，鸟嘌呤环化酶激活蛋白（GCAP）（一种对Ca2 +敏感的激活剂）以低细胞内Ca2 +浓度刺激了鸟嘌呤环化酶（RetGC），以重新合成光消耗的cGMP。在洗涤过的外段膜中，我们发现GCAP可刺激的RetGC在生理温度（30-37摄氏度）下迅速失活。在相同条件下，RetGC仍能胜任S-100蛋白制剂或Mn2 + / Triton X-100的刺激，表明环化酶催化域保持功能。 GCAP和腺嘌呤核苷酸可防止失活。 GCAP的保护与Ca2 +浓度无关，这表明GCAP与RetGC之间存在Ca2 +独立的相互作用。 ATP（EC50 = 150 microM）的保护不是由于磷酸化引起的，因为不可水解的类似物腺苷亚胺基二磷酸（AMP-PNP）的保护效果相同。除了保护作用外，ATP和AMP-PNP还可以缓慢刺激环化酶活性。与生理温度下RetGC的功能变化同时，我们还观察到结构变化。除非存在ATP，AMP-PNP或GCAP，否则在30°C下预温育后，RetGC-1的62 kDa细胞内片段对胰蛋白酶的切割变得更加敏感。因此，腺嘌呤核苷酸和GCAP在结构上和功能上都保护RetGC。

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