Mycoplasma synoviae dihydrolipoamide dehydrogenase is an immunogenic fibronectin/plasminogen binding protein and a putative adhesin
Mycoplasma synoviae dihydrolipoamide dehydrogenase is an immunogenic fibronectin/plasminogen binding protein and a putative adhesin
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滑液支原体二氢硫辛酰胺脱氢酶是一种免疫原性纤连蛋白/纤溶酶原结合蛋白和假定的粘附素
DOI：
10.1016/j.vetmic.2021.109328
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发表时间：
2022-01-12
影响因子：
3.3
通讯作者：
Yu, Shengqing
中科院分区：
文献类型：
--
作者：
Qi, Jingjing;Wang, Yu;Yu, Shengqing
滑膜支原体（Mycoplasma synoviae， M. synoviae）是一种重要的禽类病原体，可引起雏鸡和火鸡的关节炎和空气性毛囊炎。滑膜分枝杆菌感染给全世界的家禽业造成了相当大的经济损失。细胞粘附是支原体感染的关键阶段。二氢脂酰胺脱氢酶（PdhD）是一种黄素依赖性酶，对能量代谢和氧化还原平衡至关重要。迄今为止，人们对其在细胞粘附中的作用知之甚少。本研究利用大肠杆菌BL21上清成分表达滑膜分枝杆菌重组蛋白（rMSPdhD），制备兔抗rMSPdhD血清。免疫印迹实验表明，rMSPdhD是一种免疫原性蛋白，而支原体杀灭实验显示，兔抗rMSPdhD血清具有较高的补体依赖性支原体杀灭率（88.5%）。通过悬浮免疫荧光法和亚细胞定位分析，发现MSPdhD是一种表面定位蛋白，分布在滑膜分枝杆菌的细胞质和细胞膜中。rMSPdhD的酶活性通过测定其还原脂酰胺为二氢脂酰胺和将NADH转化为NAD（+）的能力来确定。通过间接免疫荧光实验，rMSPdhD可以粘附在DF-1鸡胚成纤维细胞上。兔抗rmspdhd血清能明显抑制滑膜分枝杆菌对DF-1细胞的附着。Western blot和ELISA结合实验证实，rMSPdhD还与纤维连接蛋白（Fn）和纤溶酶原（Plg）呈剂量依赖性结合。综上所述，我们的数据表明MSPdhD不仅是一种生物酶，而且是一种免疫原性表面暴露蛋白，可以与Fn和Plg结合并粘附在宿主细胞上。此外，我们发现兔抗rmspdhd血清可以抑制滑膜分枝杆菌对DF-1细胞的粘附，并具有显著的补体依赖性杀菌活性。我们的发现提示MSPdhD可能参与滑膜分枝杆菌的发病机制。
Mycoplasma synoviae (M. synoviae) is an important avian pathogen that causes arthritis and airsacculitis in young chickens and turkeys. Infection by M. synoviae results in considerable economic losses to the poultry industry worldwide. Cytoadherence is a crucial stage during mycoplasma infection. Dihydrolipoamide dehydrogenase (PdhD) is a flavin-dependent enzyme that is critical for energy metabolism and redox balance. To date, its role in cytoadherence is poorly understood. In this study, recombinant PdhD from M. synoviae (rMSPdhD) was expressed in the supernatant component of E. coli BL21 and rabbit anti-rMSPdhD serum was prepared. rMSPdhD was shown to be an immunogenic protein by immunoblot assays, while the mycoplasmacidal assay revealed that the rabbit anti-rMSPdhD serum had a high complement-dependent mycoplasmacidal rate (88.5 %). Using a suspension immunofluorescence assay and subcellular localization analysis, MSPdhD was shown to be a surface-localized protein distributed in both the cytoplasm and cell membrane of M. synoviae. The enzymatic activity of rMSPdhD was determined by measuring its ability to reduce lipoamide to dihydrolipoamide and convert NADH to NAD(+). Using an indirect immunofluorescence assay, rMSPdhD was shown to adhere to DF-1 chicken embryo fibroblast cells. Furthermore, the attachment of M. synoviae to DF-1 cells was significantly inhibited by rabbit anti-rMSPdhD serum. Western blot and ELISA binding assays confirmed that rMSPdhD also bound to fibronectin (Fn) and plasminogen (Plg) in a dose-dependent manner. In conclusion, our data show that MSPdhD is not only a biological enzyme, but also an immunogenic surface-exposed protein that can bind to Fn and Plg as well as adhere to host cells. In addition, we show that rabbit anti-rMSPdhD serum can inhibit the adhesion of M. synoviae to DF-1 cells and has a significant complement-dependent bactericidal activity. Our findings suggest that MSPdhD may be involved in the pathogenesis of M. synoviae.