Alberdi F, Alderton M R, Korolik V, Coloe P J, Smith S C
Department of Applied Biology and Biotechnology, Royal Melbourne Institute of Technology University, Victoria, Australia.
Immunol Cell Biol. 1995 Feb;73(1):38-43. doi: 10.1038/icb.1995.6.
Antibacterial peptides were purified from porcine neutrophil granules collected from healthy pigs. Granule proteins, extracted with 0.2 mol/L sodium acetate were subjected to ion-exchange chromatography and five peaks (designated A to E) were detected. Individual porcine neutrophil granule proteins were shown to inhibit the growth of target organisms Escherichia coli and Staphylococcus aureus. The antimicrobial activity was shown to be concentration and time dependent. Peak D showed strong antimicrobial activity against S. aureus and peak C (with a greater number of eluted proteins) was shown to be active against both S. aureus and E. coli. One of the peptides was purified further by reverse-phase HPLC from peak fraction C. The MW of this peptide was approximately 5500 Da as determined by SDS-PAGE and mass spectral analysis and was active against both E. coli and S. aureus in vitro sustaining a > 90% decrease, respectively, in CFU after a 2 h exposure with 50 micrograms of this peptide. Amino acid analysis showed the peptide was rich in aspartate/aspartic acid, glutamine/glutamic acid, proline, arginine and threonine. The antimicrobial activity of this peptide and other novel proteins in porcine neutrophilic granules demonstrates the probable role of these proteins and peptides in host defence of porcine neutrophils against bacterial infection.
抗菌肽是从健康猪体内采集的猪嗜中性粒细胞颗粒中纯化得到的。用0.2 mol/L醋酸钠提取的颗粒蛋白进行离子交换色谱分析，检测到五个峰（命名为A至E）。已证明单个猪嗜中性粒细胞颗粒蛋白可抑制目标生物体大肠杆菌和金黄色葡萄球菌的生长。抗菌活性呈浓度和时间依赖性。峰D对金黄色葡萄球菌显示出较强的抗菌活性，峰C（洗脱蛋白数量较多）对金黄色葡萄球菌和大肠杆菌均有活性。从峰馏分C中通过反相高效液相色谱进一步纯化了一种肽。通过SDS-PAGE和质谱分析确定该肽的分子量约为5500 Da，在体外对大肠杆菌和金黄色葡萄球菌均有活性，用50微克该肽处理2小时后，CFU分别下降>90%。氨基酸分析表明该肽富含天冬氨酸/天冬氨酸、谷氨酰胺/谷氨酸、脯氨酸、精氨酸和苏氨酸。该肽和猪嗜中性粒细胞颗粒中其他新蛋白的抗菌活性表明这些蛋白和肽在猪嗜中性粒细胞抵御细菌感染的宿主防御中可能发挥的作用。
