权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Analysis of activation factor for NADPH oxidase in neutrofhils

中性粒细胞NADPH氧化酶激活因子分析

基本信息

批准号：
04671354
负责人：
OKUMURA Naoki
金额：
$ 1.09万
依托单位：
HIROSHIMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

项目摘要

NADPH oxidase in neutrophils plays an important role in killing invaded bacteria. We found a low molecular cytosolic activation factor in guinea pig neutrophils for NADPH oxidase in a cell-free system, which consisting plasma membranes, cytosol and arachidonic acid. The cytosolic fraction was separated into a high molecular weight fraction larger than 10 kDA (a through fraction) and low molecular weight fractions by sephadex G-25 gel filtration chromatography. Inthe cell-free system, NADPH oxidase was activated using the high molecular weight fraction and each of the low molecular weight fractions instead of cytosol. The low molecular weight faction, which contained about 1kDa materials and had no ability to activate NADPH oxidase, enhanced NADPH oxidase activation by the high molecular weight fraction. Treatment of the low molecular weight faction with heat and proteinase did not affect the enhancement. In the presence of FAD and GTPsigmaS, well-known low molecular weight activation f … More actors, the low molecular weight fraction enhanced NADPH oxidase activation. These results indicate the low molecular weight fraction contains a new cytosolic activation factor for NADPH oxidase.Onthe other hand, we also investigated phosphorylation and dephosphorylation of 46 kDa protein in guinea pig neutrophils to clarify the importance of phosphorytion of this protein to NADPH oxidase actvation. In the plasma membranes prepared from activaed neutrophils, we found the correlation between dephosphrylaion of the 46 kDa protein and deactivation of NADPH oxidase. Furthermore, the deactivation of NADPH oxidase was suppressed by a protein phoshatase inhibitor. By the treatment of neutrophils with the inhibitor, a weak and temporal activation by fMLP, a chemotactic peptide, changed to substantial and prolonged activation. These reslts shows that the dephosphorylation of 46 kDa protein cause the deactivation of NADPH oxidase, and not only the increase in the phosphorylation of 46 kDa protein but also the decrease in the dephosohorylation of the protein activate NADPH oxidase. Less

中性粒细胞中的NADPH氧化酶在杀死入侵细菌中起重要作用。我们在豚鼠中性粒细胞中发现了一种低分子胞浆活化因子，用于NADPH氧化酶的无细胞系统，该系统由质膜、胞浆和花生四烯酸组成。采用sephadex G-25凝胶过滤层析法将胞质组分分离为大于10 kDA的高分子量组分（a through组分）和低分子量组分。在无细胞系统中，用高分子量组分和每一个低分子量组分代替细胞质溶胶激活NADPH氧化酶。低分子量部分含有约1kDa的物质，没有激活NADPH氧化酶的能力，而高分子量部分则增强了NADPH氧化酶的激活。热处理和蛋白酶处理对低分子量组分的增强效果没有影响。在FAD和GTPsigmaS存在的情况下，众所周知的低分子量激活作用，更多的参与者，低分子量部分增强了NADPH氧化酶的激活。这些结果表明，低分子量组分含有一种新的NADPH氧化酶胞浆活化因子。另一方面，我们还研究了豚鼠中性粒细胞中46 kDa蛋白的磷酸化和去磷酸化，以阐明该蛋白的磷酸化对NADPH氧化酶激活的重要性。在活化的中性粒细胞制备的质膜中，我们发现46 kDa蛋白的去磷酸化与NADPH氧化酶的失活之间存在相关性。此外，NADPH氧化酶的失活被一种蛋白磷酸酶抑制剂抑制。通过用抑制剂处理中性粒细胞，fMLP（一种趋化肽）的微弱和短暂激活转变为实质性和长期激活。这些结果表明，46 kDa蛋白的去磷酸化引起NADPH氧化酶失活，并且46 kDa蛋白的磷酸化水平升高，同时该蛋白的去磷酸化水平降低，从而激活NADPH氧化酶。少