人LAK细胞免疫效应分子HMGN2的鉴定

Identification of HMGN2 as an Effector Molecule of Human LAK Cells

为分离纯化人淋巴因子激活的杀伤细胞(LAK)小分子抗菌多肽,应用制备尿素-聚丙烯酰胺凝胶电泳技术和反向高效液相色谱技术分离纯化人LAK细胞酸溶性提取物,纯化出一个具抗菌活性的多肽HLP-3p21.蛋白质N端氨基酸测序、质谱精确分子质量测定、蛋白质印迹分析证明HLP-3p21为HMGN2.最小抑菌浓度(MIC)和最小杀菌浓度(MBC)试验证明HMGN2有抗大肠杆菌ML-35p氨苄青霉素耐药株、铜绿假单胞菌ATCC27853、白色念珠菌ATCC10231活性,无抗金黄色葡萄球菌ATCC25923活性.制备HMGN2多克隆抗体,应用免疫荧光化学、酶联免疫吸附测定和蛋白质印迹方法对HMGN2进行定位分析,证明单个核细胞经IL-2刺激成为LAK细胞时部分HMGN2由胞核转移至胞浆,进而分泌到胞外.提示HMGN2是LAK细胞一个新的免疫效应分子.

An antimicrobial polypeptide was isolated and purified from the acid soluble proteins of human LAK cells. Its N-terminal amino sequence was identical to HMGN2 (high mobility group nucleosomal binding domain 2). Mass spectrum identification and Western blotting analysis also indicated its individual character of HMGN2. The antimicrobial assay showed that MICs of the recombinant HMGN2 against E.coli ML-35p (an ampiciline-resistance strain), Pseudomonas aeruginosa ATCC 27853, and Candida albicans ATCC 10231 were 12.5, 25, and 100 mg/L respectively. In contrast, the recombinant holo-HA4G-17 was inactive against Staphylococcus aureus ATCC 25923. The immunocytochemistry staining, ELISA, and Western blotting revealed that HMGN2 was present in the cytoplasm of mononuclear leukocytes and released to the extracellular environment when stimulated with IL-2. The results indicated that HMGN2 was a new effector molecule of human LAK cells.