Institute of Medical Microbiology, Immunology and ParasitologyPharmaceutical Microbiology Section, University of Bonn,53105 Bonn, Germany.
Infect Immun. 2010 Jun;78(6):2793-800. doi: 10.1128/IAI.00688-09. Epub 2010 Apr 12.
Human beta-defensin 3 (hBD3) is a highly charged (+11) cationic host defense peptide, produced by epithelial cells and neutrophils. hBD3 retains antimicrobial activity against a broad range of pathogens, including multiresistant Staphylococcus aureus, even under high-salt conditions. Whereas antimicrobial host defense peptides are assumed to act by permeabilizing cell membranes, the transcriptional response pattern of hBD3-treated staphylococcal cells resembled that of vancomycin-treated cells (V. Sass, U. Pag, A. Tossi, G. Bierbaum, and H. G. Sahl, Int. J. Med. Microbiol. 298:619-633, 2008) and suggested that inhibition of cell wall biosynthesis is a major component of the killing process. hBD3-treated cells, inspected by transmission electron microscopy, showed localized protrusions of cytoplasmic contents, and analysis of the intracellular pool of nucleotide-activated cell wall precursors demonstrated accumulation of the final soluble precursor, UDP-MurNAc-pentapeptide. Accumulation is typically induced by antibiotics that inhibit membrane-bound steps of cell wall biosynthesis and also demonstrates that hBD3 does not impair the biosynthetic capacity of cells and does not cause gross leakage of small cytoplasmic compounds. In in vitro assays of individual membrane-associated cell wall biosynthesis reactions (MraY, MurG, FemX, and penicillin-binding protein 2 [PBP2]), hBD3 inhibited those enzymes which use the bactoprenol-bound cell wall building block lipid II as a substrate; quantitative analysis suggested that hBD3 may stoichiometrically bind to lipid II. We report that binding of hBD3 to defined, lipid II-rich sites of cell wall biosynthesis may lead to perturbation of the biosynthesis machinery, resulting in localized lesions in the cell wall as demonstrated by electron microscopy. The lesions may then allow for osmotic rupture of cells when defensins are tested under low-salt conditions.
人β防御素 3（hBD3）是一种带正电荷（+11）的阳离子性宿主防御肽，由上皮细胞和中性粒细胞产生。hBD3 对广泛的病原体保持抗菌活性，包括耐多药金黄色葡萄球菌，即使在高盐条件下也是如此。虽然抗菌宿主防御肽被认为通过破坏细胞膜起作用，但 hBD3 处理的葡萄球菌细胞的转录反应模式类似于万古霉素处理的细胞（V. Sass、U. Pag、A. Tossi、G. Bierbaum 和 H. G. Sahl，Int. J. Med. Microbiol. 298:619-633, 2008），并表明细胞壁生物合成的抑制是杀伤过程的主要组成部分。透射电子显微镜检查 hBD3 处理的细胞显示细胞质内容物的局部突起，并且对细胞内核苷酸激活细胞壁前体的池进行分析表明最终可溶性前体 UDP-MurNAc-五肽的积累。这种积累通常是由抑制细胞壁生物合成膜结合步骤的抗生素诱导的，并且还表明 hBD3 不会损害细胞的生物合成能力，并且不会导致细胞质小分子的大量泄漏。在单个膜相关细胞壁生物合成反应（MraY、MurG、FemX 和青霉素结合蛋白 2 [PBP2]）的体外测定中，hBD3 抑制了那些使用 bactoprenol 结合细胞壁构建块脂质 II 作为底物的酶；定量分析表明 hBD3 可能与脂质 II 成化学计量结合。我们报告 hBD3 与细胞壁生物合成的定义的、富含脂质 II 的位点结合可能导致生物合成机制的扰动，导致电子显微镜下细胞壁的局部损伤。然后，当在低盐条件下测试防御素时，这些损伤可能允许细胞通过渗透破裂。
