Lysozymes, which are secreted in many organisms, including invertebrates, mammals, plants, bacteria and fungus, exhibit antimicrobial, antiviral, antioxidant, and anti-inflammatory activities. Splys-i is an invertebrate-type (i-type) lysozyme isolated from Scylla paramamosain in 2017 and is involved in immune defense against bacteria. However, the antibacterial, antioxidant, and anti-inflammatory activities of Splys-i remain to be elucidated. In the current study, the expression parameters (including IPTG concentration, induction temperature, and induction duration) of Splys-i in Escherichia coli were optimized to achieve high-level yield through shake-flask cultivation with approximately 120 mg of Splys-i obtained from 1 L of LB medium. The purified Splys-i displayed low cytotoxicity to RAW264.7 macrophage cells and low hemolytic activity against erythrocytes of mouse, rat, and rabbit, respectively, and exhibited potent antibacterial activity against both Gram-positive and -negative bacteria with minimum concentrations ranging from 15 to 90 μg/mL. The antibacterial property of Splys-i was also unaffected when treated with various temperature, pHs, and salinity, respectively, and Splys-i showed resistance to proteinase digestion. Radical-scavenging rate assay (including ABTS⁺, DPPH, hydroyl free radical, and superoxide anion) indicated that Splys-i was an efficient antioxidant. Splys-i also exerted anti-inflammatory effect through the inhibition of IκBα and NF-κB(P65) phosphorylation, thereby reducing the secretion of pro-inflammatory cytokines. All these results suggested that Splys-i can be prepared from E. coli with potent biological property.

在许多生物（包括无脊椎动物，哺乳动物，植物，细菌和真菌）中分泌的溶菌酶具有抗微生物，抗病毒，抗氧化剂和抗炎活性。Splys-i是一种无脊椎动物型（i-型）溶菌酶，于2017年从Scylla paramamosain中分离出来，参与了针对细菌的免疫防御。但是，Splys-i的抗菌，抗氧化剂和抗炎活性仍有待阐明。在本研究中，Splys-i在大肠杆菌中的表达参数（包括IPTG浓度，诱导温度和诱导持续时间）用从1 L LB培养基中获得的约120 mg Splys-i进行摇瓶培养，优化大豆蛋白以获得高产量。纯化的Splys-i分别对RAW264.7巨噬细胞具有较低的细胞毒性，并且对小鼠，大鼠和兔子的红细胞均具有较低的溶血活性，并且对革兰氏阳性和阴性细菌均具有有效的抗菌活性，其最低浓度范围为15至90μg/ mL。当分别用各种温度，pH和盐度处理时，Splys-i的抗菌性能也不受影响，并且Splys-i显示出对蛋白酶消化的抵抗力。自由基清除率测定法（包括ABTS ⁺，DPPH，氢氧自由基和超氧阴离子）表明Splys-i是有效的抗氧化剂。Splys-i还通过抑制IκBα和NF-κB（P65）磷酸化发挥抗炎作用，从而减少了促炎细胞因子的分泌。所有这些结果表明，可以从具有强生物学特性的大肠杆菌制备Splys-i 。