The peptide-binding cavity is essential for Als3-mediated adhesion of Candida albicans to human cells.
The peptide-binding cavity is essential for Als3-mediated adhesion of Candida albicans to human cells.
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DOI：
10.1074/jbc.m114.547877
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发表时间：
2014-06-27
期刊：
影响因子：
--
通讯作者：
Cota E
中科院分区：
文献类型：
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作者：
Lin J;Oh SH;Jones R;Garnett JA;Salgado PS;Rusnakova S;Matthews SJ;Hoyer LL;Cota E
背景：在C.白念珠菌Als家族中，Als 3对人类细胞粘附的贡献最大。结果：Als 3蛋白结合腔（PBC）的突变导致Als 3粘附功能的丧失。结论：PBC是Als 3粘附功能的必要条件。意义：干扰PBC功能是抑制C。白色念珠菌粘连白色念珠菌的粘附表型有助于其定殖宿主并引起疾病的能力。Als蛋白是研究最广泛的C.白色念珠菌毒力属性; ALS 3的缺失产生粘附功能的最大降低。虽然粘附活性被认为存在于Als蛋白（NT-Als）的N-末端结构域内，但粘附的分子机制仍不清楚。我们设计了NT-Als 3中的突变，以测试肽结合腔（PBC）对C的贡献。白色念珠菌粘附，并评估了在不存在功能性PBC的情况下其他NT-Als 3特征的粘附特性。纯化的PBC功能丧失突变蛋白的结构分析表明，突变不改变NT-Als 3的整体结构或表面性质。将突变掺入全长ALS 3中，并在天然ALS 3启动子的控制下整合到缺失突变体的ALS 3基因座中。PBC突变体表型在使用单层人咽上皮和脐静脉内皮细胞以及新鲜收集的人颊上皮细胞悬液的测定中进行评价。PBC功能丧失导致与Δ als 3/Δ als 3菌株无法区分的粘附表型。还使用这些方法测试了Als 3淀粉样蛋白形成区（AFR）的粘附贡献。C.白念珠菌菌株生产的细胞表面Als 3中，淀粉样蛋白的潜力被破坏的AFR粘附的贡献很小，而不是建议的AFR的聚集功能。总的来说，这些结果表明PBC在Als 3粘附的基本和主要作用。
Background: Of the eight cell surface glycoproteins in the C. albicans Als family, Als3 makes the largest contribution to adhesion to human cells. Results: Mutation of the Als3 peptide-binding cavity (PBC) results in loss of Als3 adhesive function. Conclusion: The PBC is required for Als3 adhesive function. Significance: Interfering with PBC function is a viable strategy for inhibiting C. albicans adhesion. The adhesive phenotype of Candida albicans contributes to its ability to colonize the host and cause disease. Als proteins are one of the most widely studied C. albicans virulence attributes; deletion of ALS3 produces the greatest reduction in adhesive function. Although adhesive activity is thought to reside within the N-terminal domain of Als proteins (NT-Als), the molecular mechanism of adhesion remains unclear. We designed mutations in NT-Als3 that test the contribution of the peptide-binding cavity (PBC) to C. albicans adhesion and assessed the adhesive properties of other NT-Als3 features in the absence of a functional PBC. Structural analysis of purified loss-of-PBC-function mutant proteins showed that the mutations did not alter the overall structure or surface properties of NT-Als3. The mutations were incorporated into full-length ALS3 and integrated into the ALS3 locus of a deletion mutant, under control of the native ALS3 promoter. The PBC mutant phenotype was evaluated in assays using monolayers of human pharyngeal epithelial and umbilical vein endothelial cells, and freshly collected human buccal epithelial cells in suspension. Loss of PBC function resulted in an adhesion phenotype that was indistinguishable from the Δals3/Δals3 strain. The adhesive contribution of the Als3 amyloid-forming-region (AFR) was also tested using these methods. C. albicans strains producing cell surface Als3 in which the amyloidogenic potential was destroyed showed little contribution of the AFR to adhesion, instead suggesting an aggregative function for the AFR. Collectively, these results demonstrate the essential and principal role of the PBC in Als3 adhesion.