葡萄球菌和链球菌噬菌体裂解酶嵌合蛋白的构建与作用机制研究

Pathogenic bacteria infection is the main reason of bovine mastitis. Due to bacterial drug resistance, antibiotic-resistant strains increased, lead to poor therapeutic effect of antibiotic treatment. Bacteriophage lysins can efficient lytic bacteria, not infect eukaryotic cells, also not destroy the body's normal flora, and have completely different action mechanism of antibiotics. So, bacteriophage lysins have potential as a new type of antibacterial drug to treat bovine mastitis. Bovine mastitis caused by pathogens, especially staphylococci, streptococcal, and Escherichia coli infection. Bacteriophage lysins with high specificity has certain limitation in the bovine mastitis treatment. Thus, in the following work, we constructed chimeric protein by lytic domain from lysins encoded by staphylococci and streptococcal bacteriophages. More importantly, it is influence the chimeric protein activity， spatial conformation and interaction relationship between in different activity domain. For further study the protective effect of chimeric protein on BMECs， the effects of chimeric protein on milk fat and milk protein pretherapy and post-treatment，the antigenicity and the pharmacokinetic of chimeric protein. These studies will provide a reference for chimeric protein as a new type of antibacterial drug to treat bovine mastitis.

病原菌感染是导致奶牛乳房炎主要病因，由于细菌耐药性不断增强、耐药菌株大量出现，导致抗生素疗效不佳。噬菌体裂解酶可以高效的裂解细菌，不侵染真核细胞，也不会破坏机体的正常菌群，有着与抗生素完全不同的作用机制。因此，噬菌体裂解酶有潜力作为新型抗菌药物治疗奶牛乳房炎。奶牛乳房炎病原菌种类多，尤其是葡萄球菌、链球菌和大肠杆菌感染。裂解酶又具有较高的特异性，因而在治疗奶牛乳房炎时具有一定的局限性。本项目拟构建奶牛乳房炎主要致病的葡萄球菌和链球菌噬菌体裂解酶裂解活性区域嵌合蛋白，重点研究嵌合蛋白活性、空间构象的影响以及嵌合蛋白内不同活性区域之间相互作用关系；进一步研究嵌合蛋白对奶牛乳腺上皮细胞的保护作用及治疗前后对乳脂、乳蛋白的影响，研究嵌合蛋白进入体内后的抗原性、与组织细胞的相互作用及其体内代谢规律等特性，为将噬菌体裂解酶嵌合蛋白作为新型抗菌药物治疗奶牛乳房炎或其它细菌感染提供参考。

病原菌感染是导致奶牛乳房炎主要病因，由于细菌耐药性不断增强、耐药菌株大量出现，导致抗生素疗效不佳。噬菌体裂解酶可以高效的裂解细菌，不侵染真核细胞，也不会破坏机体的正常菌群，有着与抗生素完全不同的作用机制。在国家自然科学基金青年基金项目资助下，项目组成功分离获得金黄色葡萄球菌噬菌体，测定生物学特性及裂解活性；建立不同材质细菌生物被膜模型，利用模型评价噬菌体对不同材质生物被膜的清除作用效果；对噬菌体全基因组序列测定、生物信息学分析确定裂解酶裂解域，对其进行表达及抗金黄色葡萄球菌和无乳链球菌活性测定；确定白屈菜红碱对金黄色葡萄球菌和无乳链球菌抗菌活性和作用机制。上述研究为将噬菌体裂解酶结构域作为新型抗菌药物治疗奶牛乳房炎或其它细菌感染提供参考。