Zhang Yue, Lin Hong, Wang Jingxue, Li Mengzhe
Food Safety Laboratory, Department of Food Science and Engineering, Ocean University of China, Qingdao 266003, People's Republic of China.
J Food Prot. 2018 Feb;81(2):332-340. doi: 10.4315/0362-028X.JFP-17-144.
Although Shewanella putrefaciens is the specific spoilage organism in most seafood, only seven Shewanella phages have been sequenced and their endolysins have not been reported until now. In this study, we cloned and expressed two lysis-related proteins (Spp64 and Spp62) encoded by phage Spp001, the first sequenced S. putrefaciens phage. Both recombinant proteins showed strong lytic capability toward chilled S. putrefaciens Sp225 and presented a wider activity spectrum compared with bacteriophage Spp001. The enzymatic activity of crude Spp64, Spp62, and Spp62-GST can cause decreases of 0.691, 0.674, and 0.685, respectively, as tested through the turbidity reduction assay. Furthermore, purified enzyme Spp64 at concentrations of 537.5 and 4.20 μg/mL was enough to decrease the optical density of chilled S. putrefaciens by 0.881 and 0.492, respectively, within 15 min. The recombinant Spp64 has a peptidase catalytic domain and exhibits high temperature resistance. Moreover, Spp64 displayed superior enzymatic activity in a range of pH values that matches environmental conditions (pH between 5.0 and 10.0), which demonstrates that its application in seafood is feasible. The present work is to our knowledge the first report on lysis-related enzymes encoded in the Shewanella phage. Both proteins presented extraordinary potential to control S. putrefaciens; we hope that these proteins can be developed as novel antibacterial agents in further research.
尽管腐败希瓦氏菌是大多数海产品中的特定腐败菌，但目前仅对7种希瓦氏菌噬菌体进行了测序，其溶菌酶尚未见报道。在本研究中，我们克隆并表达了噬菌体Spp001（首个测序的腐败希瓦氏菌噬菌体）编码的两种与裂解相关的蛋白（Spp64和Spp62）。两种重组蛋白对冷藏的腐败希瓦氏菌Sp225均表现出强大的裂解能力，且与噬菌体Spp001相比具有更宽的活性谱。通过浊度降低试验测试，粗提的Spp64、Spp62和Spp62 - GST的酶活性分别可使吸光度降低0.691、0.674和0.685。此外，浓度为537.5和4.20 μg/mL的纯化酶Spp64在15分钟内分别足以使冷藏的腐败希瓦氏菌的光密度降低0.881和0.492。重组Spp64具有肽酶催化结构域并表现出耐高温性。此外，Spp64在一系列与环境条件匹配的pH值（pH在5.0至10.0之间）范围内表现出优异的酶活性，这表明其在海产品中的应用是可行的。据我们所知，本研究是关于希瓦氏菌噬菌体中编码的与裂解相关酶的首次报道。这两种蛋白在控制腐败希瓦氏菌方面均展现出巨大潜力；我们希望这些蛋白能在进一步研究中开发成为新型抗菌剂。
