Rao S S, Kochhar S, Kochhar V K
National Botanical Research Institute, Lucknow, India.
Biochem Mol Biol Int. 1999 Mar;47(3):347-60. doi: 10.1080/15216549900201373.
Aspartate kinase (AK) activity is regulated by light. The activity was more in light exposed barley seedlings than those grown in the dark. The light effect was manifested even with small exposures of 5 min duration and red light was more effective than white light in this respect. The effect of 5 min red light could be reversed by a 5 min pulse of far-red light indicating the involvement of phytochrome in this response. The phytochrome is also involved in long term light effects (24 hr exposures with white light). Ca++ takes part in the signal transduction pathway for this light response. Western blot analysis using antibodies raised against the purified lysine- and threonine-sensitive AK isoenzymes from spinach leaves showed no cross reaction with the antibodies to the threonine-sensitive AK in the dark and 5 min far-red light exposed seedlings. But the protein band was detected in the white and red lights. Northern blot analysis of seedlings grown under dark and exposed to white, red and far-red lights and probed with the gene encoding aspartokinase-homoserine dehydrogenase (AKHSD) protein indicated that the gene was differentially expressed. In dark grown seedlings, AKHSD transcript was in low concentration as compared to white light where the transcript concentration was high. A 5 min red light pulse increased the transcript concentration significantly in contrast to 5 min far-red light. The transcript concentration was reduced when 5 min red light was followed by a 5 min far-red light pulse. The AK activity in dark-raised seedlings is attributed to the presence of only one isoenzyme that is sensitive to lysine but insensitive to Ca++ and calmodulin (CAM). In both white and red light exposed seedlings, three isoenzymes of AK were detected. Two of these were sensitive to threonine while one was sensitive to lysine. Both of the threonine sensitive isoenzymes were activated by Ca++ and CAM. Also one of these isoenzymes seems to be located and synthesized in chloroplasts because its synthesis was completely inhibited by chloramphenicol but not by cycloheximide.
天冬氨酸激酶（AK）的活性受光照调节。暴露在光照下的大麦幼苗中的该活性比在黑暗中生长的幼苗更高。即使光照时间短至5分钟，光照效应也会显现，在这方面，红光比白光更有效。5分钟红光的效应可被5分钟远红光脉冲逆转，这表明光敏色素参与了此反应。光敏色素也参与长期光照效应（24小时白光照射）。Ca++参与了此光照反应的信号转导途径。使用针对从菠菜叶中纯化的赖氨酸和苏氨酸敏感型AK同工酶产生的抗体进行的蛋白质印迹分析表明，在黑暗和暴露于5分钟远红光的幼苗中，针对苏氨酸敏感型AK的抗体没有交叉反应。但在白光和红光下检测到了蛋白条带。对在黑暗中生长并暴露于白光、红光和远红光下的幼苗进行Northern印迹分析，并用编码天冬氨酸激酶-高丝氨酸脱氢酶（AKHSD）蛋白的基因进行探针检测，结果表明该基因存在差异表达。在黑暗中生长的幼苗中，AKHSD转录本浓度较低，而在白光下转录本浓度较高。与5分钟远红光相比，5分钟红光脉冲显著增加了转录本浓度。当5分钟红光后接5分钟远红光脉冲时，转录本浓度降低。黑暗培养的幼苗中的AK活性归因于仅存在一种对赖氨酸敏感但对Ca++和钙调蛋白（CAM）不敏感的同工酶。在暴露于白光和红光的幼苗中，检测到了三种AK同工酶。其中两种对苏氨酸敏感，一种对赖氨酸敏感。两种对苏氨酸敏感的同工酶均被Ca++和CAM激活。此外，其中一种同工酶似乎位于叶绿体中并在叶绿体中合成，因为其合成完全被氯霉素抑制，而不被环己酰亚胺抑制。
