PDF(Pubmed)
Abstract：
Mannheimia haemolytica is the main etiological bacterial agent in ruminant respiratory disease. M. haemolytica secretes leukotoxin, lipopolysaccharides, and proteases, which may be targeted to treat infections. We recently reported the purification and in vivo detection of a 110 kDa Zn metalloprotease with collagenase activity (110-Mh metalloprotease) in a sheep with mannheimiosis, and this protease may be an important virulence factor. Due to the increase in the number of multidrug-resistant strains of M. haemolytica, new alternatives to antibiotics are being explored; one option is lactoferrin (Lf), which is a multifunctional iron-binding glycoprotein from the innate immune system of mammals. Bovine apo-lactoferrin (apo-bLf) possesses many properties, and its bactericidal and bacteriostatic effects have been highlighted. The present study was conducted to investigate whether apo-bLf inhibits the secretion and proteolytic activity of the 110-Mh metalloprotease. This enzyme was purified and sublethal doses of apo-bLf were added to cultures of M. haemolytica or co-incubated with the 110-Mh metalloprotease. The collagenase activity was evaluated using zymography and azocoll assays. Our results showed that apo-bLf inhibited the secretion and activity of the 110-Mh metalloprotease. Molecular docking and overlay assays showed that apo-bLf bound near the active site of the 110-Mh metalloprotease, which affected its enzymatic activity.
摘要：
溶血曼海姆菌是反刍动物呼吸道疾病的主要病原。溶血M.分泌白细胞毒素，脂多糖，和蛋白酶，这可能是治疗感染的目标。我们最近报道了具有胶原酶活性的110kDaZn金属蛋白酶（110-Mh金属蛋白酶）的纯化和体内检测。这种蛋白酶可能是一种重要的毒力因子。由于溶血分枝杆菌多药耐药菌株数量的增加，正在探索抗生素的新替代品；一种选择是乳铁蛋白(Lf)，它是一种来自哺乳动物先天免疫系统的多功能铁结合糖蛋白。牛乳铁蛋白（apo-bLf）具有许多性质，其杀菌作用和抑菌作用得到了强调。进行本研究以研究apo-bLf是否抑制110-Mh金属蛋白酶的分泌和蛋白水解活性。纯化该酶，并将亚致死剂量的apo-bLf添加到溶血支原体的培养物中或与110-Mh金属蛋白酶共孵育。胶原酶活性使用酶谱和Azocoll测定进行评估。我们的结果表明，apo-bLf抑制了110-Mh金属蛋白酶的分泌和活性。分子对接和覆盖实验表明，apo-bLf结合在110-Mh金属蛋白酶的活性位点附近，这影响了它的酶活性。
