| With the development of aquacμLture, water pollution become more andmore serious. Various diseases (bacteria, viruses, parasites) appeared infishery restrict fishery. The application of plentifμL antibiotics andchemicals in prevention and cure against bacteria makes a bad affect onfish' quality, furthermore, it does harm to customers. In order toenhance production of fishery, produce healthy food of fish, it'snecessary to make a study and produce natural new antibacterial medicine.Bio-antibacterial proteins are a kind of proteins with antibacterialactivity, which exist in organisms widely and play an important role inresisting pathogens. They are looked as the significant component forrecovery in organisms which are lack of adaptive immunity. Silurusmeridionalis and Spinibarbus sinensis, whose flesh are fresh anddelicious, grow speedily, so the market of them is vast. It will improvethe economy that research with the antibacterial-proteins andantibacterial-mechanism.Silurus meridionalis and Spinibarbus sinensis, were got from TongWeiAquatic Science & Technology Co., Ltd.In order to study antibacterial proteins, we first made a study onthe agglutinin isolated from serum of Silurus meridionalis and Spinibarbus sinensis. Then we researched antibacterial activity andcharacterization of antibacterial-agent extracted with three systemsseparately from skin secretion and other tissues of both healthy anddiseased Silurus meridionalis.A natural occurring agglutinin, which was detected from serum ofSilurus meridionalis, was found to have the ability of agglutinatingdifferent kinds of antigen. The agglutinin can't permeate semipermeablefilm, it's a substance with a large molecμLar weight; treated withprotease K, Pepsin and Trypsin, the agglutinin lost agglutinatingactivity, which suggested that it's a protein; However, treated withDNase,RNase and aether, it keeped agglutinating activity unchanged,which showed that it's composition has nothing to do with nucleic acidand lipids. Incubated for 1 hour under 75℃, agglutinating activity wasunchanged; and it's not sensitive to SDS. In range from 4℃to 37℃,its agglutinating activity increases with the increment of temperature.The agglutinating activity coμLd be inhibited by the splitting of sugarand gμLactose, sucrose respectively; the resμLts demonstrated the sugarreferred above played an important role in agglutinating reaction.A natural occurring agglutinin, which was detected from serum ofSpinibarbus sinensis, was found to have the ability of agglutinatingdifferent kinds of antigen. The agglutinin can't permeate semipermeablefilm, it's a substance with a large molecμLar weight; treated withprotease K, Pepsin and Trypsin, the agglutinin lost agglutinatingactivity, which suggested that it's a protein; However, treated withDNase,RNase and aether, it keeped agglutinating activity unchanged,which showed that it's composition has nothing to do with nucleic acidand lipids. Incubated for 1 hour under 60℃, agglutinating activity wasunchanged; and it's not sensitive to SDS. In range From 4℃to 37℃, its agglutinating activity increases with the increment of temperature.The agglutinating activity coμLd be inhibited by the splitting of lactoseand sucrose respectively; the resμLts demonstrated the sugar referredabove played an important role in agglutinating reaction.With systemâ… "0.1ï¼…TritonX-100+0.2ï¼…NaAc+1mmol/L PMSF" and systemâ…¡" 0.1ï¼…TFA+50ï¼…ethanol+1mmol/L PMSF", we studied theantibacterial-agent in skin secretion,skin,stomach,intestine,liverand kidney of a healthy Silurus meridionalis, The resμLt revealed thatthe antibacterial agent extracted with systemâ…¡from skin secretionshowed most strong activity. It displayed broad-spectrum antimicrobialactivity, resisted the growth of G+ bacteria(Bacillus subtilis,Staphylococcus aureus,Staphylococcus aureus A309,Streptococcicosisfaecalis) and G- bacteria(Escherichia coli,Pseudomonas aeruginosa)strongly; Incubated with pepsin under 37℃for 30min, the antimicrobialagent lost its activity; the antimicrobial protein is stable at pH6.0or 7.0, however, the activity weakened a little at pH5 or 8; incubatedunder 100℃for 10min, the activity of supernatant was unchanged; Theantimicrobial protein was partly purified using 20％～50ï¼…Saturationdegree of (NH4)2SO4。We made a primary study about the antibacterial activity and proteinscomposition of sample extracted with systemâ…¢"10 mmol/L HEPES+112mmol/L NaCl+3.4mmol/L KCl+2.4 mmol/L NaHCO3, pH 7.4" from skin,kidney,liver of a diseased Silurus meridionalis. Agglutinin is an importantantibacterial-related protein of fish, so the main study is on the agglutininisolated from Serum of Silurus meridionalis and Spinibarbus sinensis, as made abase for further research of gene.All of these researches made a base for the next researches. |
