Biermann B J, Pao L I, Feldman L J
Department of Plant Biology, University of California, Berkeley 94720, USA.
Plant Physiol. 1994 May;105(1):243-51. doi: 10.1104/pp.105.1.243.
Protein kinase activity has repeatedly been found to co-purify with the plant photoreceptor phytochrome, suggesting that light signals received by phytochrome may be transduced or modulated through protein phosphorylation. In this study immunoprecipitation techniques were used to characterize protein kinase activity associated with phytochrome from maize (Zea mays L.). A protein kinase that specifically phosphorylated phytochrome was present in washed anti-phytochrome immunoprecipitates of etiolated coleoptile proteins. No other substrate tested was phosphorylated by this kinase. Adding salts or detergents to disrupt low-affinity protein interactions reduced background phosphorylation in immunoprecipitates without affecting phytochrome phosphorylation, indicating that the protein kinase catalytic activity is either intrinsic to the phytochrome molecule or associated with it by high-affinity interactions. Red irradiation (of coleoptiles or extracts) sufficient to approach photoconversion saturation reduced phosphorylation of immunoprecipitated phytochrome. Subsequent far-red irradiation reversed the red-light effect. Phytochrome phosphorylation was stimulated about 10-fold by a co-immunoprecipitated factor. The stimulatory factor was highest in immunoprecipitates when Mg2+ was present in immunoprecipitation reactions but remained in the supernatant in the absence of Mg2+. These observations provide strong support for the hypothesis that phytochrome-associated protein kinase modulates light responses in vivo. Since only phytochrome was found to be phosphorylated, the co-immunoprecipitated protein kinase may function to regulate receptor activity.
人们多次发现蛋白激酶活性与植物光受体光敏色素共同纯化，这表明光敏色素接收到的光信号可能通过蛋白质磷酸化进行转导或调节。在本研究中，采用免疫沉淀技术来表征与玉米（Zea mays L.）光敏色素相关的蛋白激酶活性。在黄化胚芽鞘蛋白的洗涤抗光敏色素免疫沉淀物中存在一种特异性磷酸化光敏色素的蛋白激酶。该激酶不会磷酸化其他任何测试的底物。添加盐或去污剂以破坏低亲和力蛋白质相互作用可降低免疫沉淀物中的背景磷酸化，而不影响光敏色素的磷酸化，这表明蛋白激酶催化活性要么是光敏色素分子固有的，要么通过高亲和力相互作用与其相关联。足以接近光转化饱和的红色照射（对胚芽鞘或提取物）会降低免疫沉淀的光敏色素的磷酸化。随后的远红光照射可逆转红光效应。共免疫沉淀因子可使光敏色素磷酸化增加约10倍。当免疫沉淀反应中存在Mg2+时，刺激因子在免疫沉淀物中最高，但在不存在Mg2+时仍留在上清液中。这些观察结果为光敏色素相关蛋白激酶在体内调节光反应的假说提供了有力支持。由于仅发现光敏色素被磷酸化，共免疫沉淀的蛋白激酶可能起到调节受体活性的作用。
