Inhibition of Anthrax Lethal Factor by alpha-defensins

炭疽毒素由炭疽杆菌分泌的三种蛋白质组成--保护性抗原（PA）、水肿蛋白（EAE）、 因子（EF）和致死因子（LF）。LF是一种Zn 2+依赖性金属蛋白酶，其切割蛋白酶的成员。 丝裂原活化蛋白激酶激酶（MAPKK）家族，阻断MARK信号通路， 专职吞噬细胞单独的PA和LF的二元复合物，称为致死毒素或LeTx，足以 导致靶细胞死亡并杀死实验动物，表现出与 炭疽热LF作为炭疽致病的主要毒力因子之一，已成为炭疽病的重要致病因子。 药物开发的治疗靶点。许多基于小分子的LF抑制剂已经被发现。 显示能够中和细胞培养物中LeTx的细胞毒性和动物试验中LeTx的致死性， 在炭疽感染的治疗中具有潜在的治疗价值。 人中性粒细胞α-防御素（HNP）是一个不断增长的小阳离子抗菌素家族的成员， 肽，杀死广泛的微生物，如细菌，真菌和病毒，发挥重要作用， 对传染性病原体的先天免疫防御。考夫曼和同事最近发现， HNP 1非竞争性抑制LF，阻止MAPK的切割并恢复受损的MAPK LeTx处理的巨噬细胞中的信号传导（Kim等，2005年）。此外，HNP 1从小鼠巨噬细胞中拯救了 B。a/tfftrac/s诱导的细胞毒性，并且在用HNP 1 -3进行的wVo治疗中保护小鼠免于致命的细胞毒性。 LeTx的后果（Kim et al 2005）。在随后的报告中，Mayer-Scholl等人（2005年）证明， 那个B。被人中性粒细胞吞噬并在细胞内萌发的炭疽孢子可以有效地 被HNP杀害这些发现提出了一种可能性，即α-防御素可能被开发为一类新的免疫调节剂。 对抗炭疽感染的抗毒素。 尽管基于a-防御素的抗毒素对炭疽治疗具有治疗潜力， 对于α-防御素对LF的抑制仍然知之甚少。在这份提案中，我们试图破译 序列规则和结构决定因素，决定抑制LF的α-防御素，并设计 具有显著改善的抑制活性的第二代防御素分子。

Anthrax toxin consists of three proteins secreted by Bacillus anthracis - protective antigen (PA), edema factor (EF) and lethal factor (LF). LF is a Zn2+-dependent metalloprotease that cleaves members of the mitogen-activated protein kinase kinase (MAPKK) family, blocking the MARK signaling pathway in professional phagocytes. The binary complex of PA and LF alone, termed lethal toxin or LeTx, is sufficient to nduce death of target cells and kill experimental animals, manifesting the symptoms associated with anthrax. As one of the primary virulence factors in the pathogenesis of anthrax, LF has become an important therapeutic target for drug development. A number of small-molecule-based inhibitors of LF have been shown to be able to neutralize cytotoxicityof LeTx in cell cultures and lethality of LeTx in animal trials, promising potential therapeutic value in the treatment of anthrax infection. Human neutrophil a-defensins (HNPs) are members of a growing family of small cationic antimicrobial peptides that kill a broad range of microbes such as bacteria, fungi and viruses, playing an important role in the innate immune defense against infectious pathogens. Kaufmann and colleagues recently discovered that HNP1 non-competitively inhibits LF, preventing cleavage of a MAPKK and restoring impaired MAPK signaling in LeTx-treated macrophages (Kim et al., 2005). Further, HNP1 rescued murine macrophages from B. a/tfftrac/s-induced cytotoxicity, and in wVo treatment with HNP1-3 protected mice against the fatal consequences of LeTx (Kim et al 2005). In a subsequent report, Mayer-Scholl et al. (2005) demonstrated that B. anthracis spores engulfed by human neutrophils and germinated intracellularly can be effectively killed by HNPs. These findings raise the possibility that a-defensins may be developed as a novel class of antitoxins to combat anthrax infection. Despite the therapeutic potential of a-defensin-based antitoxins for anthrax treatment, the molecular basis for the inhibition of LF by a-defensins remains poorly understood. In this proposal, we seek to decipher the sequence rules and structural determinants that dictate the inhibition of LF by a-defensins, and to design second-generation defensin molecules with significantly improved inhibitory activity.