Rta2p是本课题组首次通过对耐药菌的基因表达谱芯片分析，发现的白念珠菌耐药膜蛋白。前期研究已证明，钙离子通过钙调神经磷酸酶通路上调Rta2p表达，Rta2p通过调节细胞膜内外两侧的鞘脂分布，增强细胞膜脂筏的稳定性，从而使白念珠菌对唑类药物的损伤产生更强的抵抗力，形成耐药。本课题拟运用生物信息学、基因工程、结构生物学等相关技术，进一步考察Rta2p转运鞘脂、促进耐药的重要功能区域和关键活性位点。在此基础上，通过表达纯化Rta2p重要功能结构域，来培养结晶，对其重要功能结构域进行三维结构解析，为模拟设计其抑制剂提供结构模型。同时利用其重要功能结构域，依托BIAcore分析仪，建立其抑制剂的高通量分子筛选模型，结合真菌细胞和小鼠感染模型验证，筛选Rta2p抑制剂和协同抗耐药真菌的先导化合物，为抗真菌药物的创新研究提供关键技术，也为克服真菌耐药性、防治日益严重的深部真菌感染奠定理论基础。

To clarify the mechanism of azole resistance, our previous study found that Rta2p, an integral membrane protein, which can transport sphingolipids and modulate the balance of cholesterol and sphingolipids in lipid rafts, is involved in calcineurin-mediated azole resistance. In this study，the important amino acid residues or domains of Rta2p will be forecast by bioinformatics analysis. Mutant Rta2p, with one of the important residues or domains changed, was introduced into plasmid Bes116. The mutant plasmid, wild-type plasmid and plasmid Bes116 were used to transform JXM101 (rta2Δ/Δ mutant Candida. albicans). Depending on the phenotypic alterations, including dihydrosphingosine (DHS) transformation and azole resistance, the residues or the domains which were important in DHS export and azole resistance were identified. The crystal structural characterization, the binding affinity determination, and the enzymatic activity assay of the Rta2p mutant will be conducted to confirm the importance of the residues or the domains identified above in DHS binding and export. High throughput screening model will be established to screen the inhibitor of Rta2p.