Nitric oxide synthase(NOS) like enzyme mediated nitric oxide(NO) generation by harmful red tide phytoplankton, Chattonella marina

一氧化氮合酶（NOS）样酶介导有害赤潮浮游植物、查托氏菌码头产生一氧化氮（NO）

基本信息

批准号：
18580205
负责人：
ODA Tatsuya
金额：
$ 2.45万
依托单位：
Nagasaki University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18580205/
关键词：
Red tide phytoplankton Chattonella marina reactive oxygen species nitric oxide NO synthase toxicity flagellate

项目摘要

The unicellular marine phytoplankton Chattonella marina is known to exhibit potent fish-killing activity. Previous studies have demonstrated that C. marina produces reactive oxygen species (ROS), and ROS-mediated ichthyotoxic mechanism has been postulated. In fact, several studies have demonstrated that Chattonella spp. generate reactive oxygen species (ROS). Furthermore, our studies using electron spin resonance (ESR) spectroscopy with the spin traps 5,5-dimethyl-1-pyrroline-N-oxide and N-t-butyl-a-phenylnitrone showed that C. marina generates hydroxyl radical (OH). Since ROS are generally considered to be toxic against living organisms, the ROS generated by Chattonella spp. may at least partly involve in the fish-kill mechanism. Furthermore, we have found that C. marina exhibited ROS-mediated toxic effect on a marine bacterium, Vibrio alginolyticus. In addition to Chattonella spp., it has been reported that another raphidophycean flagellate, Heterosigma akashiwo also showed ROS-media … More ted toxicity on rainbow trout. Thus, it seems likely that the production of ROS is a common feature of raphidophycean flagellates. However, the exact toxic mechanism is still controversial. In this study, we obtained evidence that C. marina produces nitric oxide (NO)under normal growth conditions. We utilized chemiluminescence (CL)reaction between NO and luminol-H_2O_2 to detect NO in C. marina cell suspensions. In this assay, significant CL was observed in C. marina in a cell-number dependent manner, and this was diminished by the addition of carboxy-PTIO, a specific NO scavenger. The NO generation by C. marina was also confirmed by a spectrophotometric assay based on the measurement of the diazo-reaction positive substances (NO_x)and by fluorometric assay using highly specific fluorescent indicator of NO. The NO level in C. marina was significantly decreased by L-NAME, a specific NO synthase (NOS)inhibitor. The addition of L-arginine resulted in the increase in NO level, whereas NaNO_2 had no effect. These results suggest that a NOS-like enzyme is mainly responsible for NO generation in C. marina. Less

众所周知，单细胞海洋植物浮游生物Chattonella码头表现出强大的杀鱼活动。先前的研究表明，C.码头会产生活性氧（ROS），并且已经假定ROS介导的鱼毒素机制。实际上，几项研究表明查顿氏菌属属。产生活性氧（ROS）。此外，我们使用电子自旋谐振（ESR）光谱的研究与自旋陷阱5,5-二甲基-1-吡咯氨酸-N-氧化物和N-T-丁基-A-苯基硝基酮表明C. c. marina会产生羟基自由基（OH）。由于通常认为ROS是对生物体的有毒，因此Chattonella spp产生的ROS。可能至少部分参与鱼类机制。此外，我们发现C.码头暴露于ROS介导的对海洋细菌，弧菌藻素水解的毒性作用。除了查顿氏菌属，据报道，另一个Raphidophycean鞭毛蛋白酶，异性Akashiwo还显示了Ros-Media…对Rainbow Trout的TED毒性更大。这似乎是ROS的产量似乎是Raphidophycean鞭毛的共同特征。但是，确切的有毒机制仍然存在争议。在这项研究中，我们获得了证据表明，玛丽娜梭菌在正常生长条件下产生一氧化氮（NO）。我们利用NO和Luminol-H_2O_2之间的化学发光（CL）反应在C.码头细胞悬浮液中检测NO。在此测定中，以细胞数为依赖性的方式观察到了明显的CL，并且通过添加羧基PTIO（一种特定的无清除剂）减少了这一点。基于重氮反应阳性的分光光度测定法，还通过分光光度测定法证实了NO生成。物质（no_x）和通过荧光测定法使用NO的高度特异性荧光指标。 L-NAME（一种特定的NO合酶（NOS）抑制剂）显着降低了码头中的NO水平。 L-精氨酸的添加导致无水平的增加，而Nano_2无效。这些结果表明，一种类似NOS的酶主要是码头中没有产生的原因。较少的