Helmholtz Centre for Infection Research, Department of Biological Systems Analysis, Inhoffenstr.7, 38124 Braunschweig, Germany.
J Biotechnol. 2012 Jan;157(1):268-77. doi: 10.1016/j.jbiotec.2011.09.017. Epub 2011 Sep 22.
Signal transduction systems comprising histidine kinases are suggested as new molecular targets of antibiotics. The important human fungal pathogen Candida albicans possesses three histidine kinases, one of which is the type III histidine kinase CaNik1, which activates the MAP kinase Hog1. We established a screening system for inhibitors of this class of histidine kinases by functional expression of the CaNIK1 gene in S. cerevisiae. This transformant was susceptible to fungicides to which the wild type strain was resistant, such as fludioxonil and ambruticin. Growth inhibition correlated with phosphorylation of Hog1 and was dependent on an intact Hog1 pathway. At the N-terminus the histidine kinase CaNik1 has four amino acid repeats of 92 amino acids each and one truncated repeat of 72 amino acids. Within these repeats we identified 9 HAMP domains with a paired structure. We constructed mutants in which one or two pairs of these domains were deleted. S. cerevisiae transformants expressing the full-length CaNIK1 showed the highest sensitivity to the fungicides, any truncation reduced the susceptibility of the transformants to the fungicides. This indicates that the HAMP domains are decisive for the mode of action of the antifungal compounds.
包含组氨酸激酶的信号转导系统被认为是抗生素的新分子靶标。重要的人类真菌病原体白色念珠菌拥有三种组氨酸激酶，其中一种是 III 型组氨酸激酶 CaNik1，它激活 MAP 激酶 Hog1。我们通过在 S. cerevisiae 中功能性表达 CaNIK1 基因建立了此类组氨酸激酶抑制剂的筛选系统。该转化体易受野生型菌株耐药的杀真菌剂（如氟啶酮和安布曲霉素）的影响。生长抑制与 Hog1 的磷酸化相关，并且依赖于完整的 Hog1 途径。在 N 端，组氨酸激酶 CaNik1 具有四个重复 92 个氨基酸的氨基酸重复序列和一个截断的重复序列 72 个氨基酸。在这些重复序列中，我们鉴定出 9 个具有配对结构的 HAMP 结构域。我们构建了缺失一个或两个对这些结构域的突变体。表达全长 CaNIK1 的 S. cerevisiae 转化体对杀真菌剂表现出最高的敏感性，任何截断都会降低转化体对杀真菌剂的敏感性。这表明 HAMP 结构域对这些抗真菌化合物的作用模式具有决定性作用。
