Marzari R, Scaggiante B, Skerlavaj B, Bittolo M, Gennaro R, Romeo D
Department of Biology, University of Trieste, Italy.
Infect Immun. 1988 Aug;56(8):2193-7. doi: 10.1128/iai.56.8.2193-2197.1988.
Monoclonal antibodies were raised against a bactericidal protein fraction that was purified from an extract of bovine neutrophil granules and that was previously shown (A. Savoini, R. Marzari, L. Dolzani, D. Serranò, G. Graziosi, R. Gennaro, and D. Romeo, Antimicrob. Agents Chemother. 26:405-407, 1984) to inhibit bacterial DNA synthesis. One of these antibodies, BP97, was covalently linked to Affi-Gel 10 and was used for immunoaffinity chromatography of granule extracts. Elution of the bound proteins, followed by reversed-phase high-performance liquid chromatography, generated several peptides whose molecular weights fell in the range of 1,600 to 64,000 and which reacted to BP97 but not to other mouse monoclonal or polyclonal antibodies. The reaction to BP97 appeared to be specific, inasmuch as a full panel of cationic oligo- or polypeptides was not recognized by this monoclonal antibody. Among the purified granule polypeptides, the more cationic ones (with molecular weights of 4,300 to 8,000) inhibited the growth of Escherichia coli at a MIC of 12 to 50 micrograms/ml. In addition, a 1,600-molecular-weight, highly cationic peptide also inhibited the growth of Staphylococcus aureus and Staphylococcus epidermidis at MICs of 3 to 8 micrograms/ml.
制备了针对一种杀菌蛋白组分的单克隆抗体，该组分从牛中性粒细胞颗粒提取物中纯化得到，先前已证明（A. 萨沃伊尼、R. 马尔扎里、L. 多尔扎尼、D. 塞拉诺、G. 格拉齐奥西、R. 热纳罗和D. 罗密欧，《抗菌剂与化疗》26:405 - 407，1984年）其可抑制细菌DNA合成。其中一种抗体BP97与Affi - Gel 10共价连接，并用于颗粒提取物的免疫亲和层析。对结合蛋白进行洗脱，随后进行反相高效液相色谱分析，得到了几种肽，其分子量在1600至64000范围内，与BP97反应，但不与其他小鼠单克隆或多克隆抗体反应。对BP97的反应似乎具有特异性，因为该单克隆抗体不能识别一整套阳离子寡肽或多肽。在纯化的颗粒多肽中，阳离子性更强的那些（分子量为4300至8000）在12至50微克/毫升的最低抑菌浓度下可抑制大肠杆菌的生长。此外，一种分子量为1600的高度阳离子性肽在3至8微克/毫升的最低抑菌浓度下也可抑制金黄色葡萄球菌和表皮葡萄球菌的生长。
