Saxena S, Madan T, Muralidhar K, Sarma P U
Institute of Genomics and Integrative Biology, Delhi, India.
Clin Exp Immunol. 2003 Oct;134(1):86-91. doi: 10.1046/j.1365-2249.2003.02257.x.
Aspergillus fumigatus (Afu) is an important fungal pathogen causing allergic and invasive respiratory disorders. A plethora of multi-functional allergens/antigens secreted by Afu have been implicated in pathogenesis. The present study was undertaken to identify and characterize novel Afu allergen/antigen by cDNA library approach. cDNA library of Afu was immunoscreened with pooled sera of allergic bronchopulmonary aspergillosis (ABPA) patients. The cDNA clone, TS1, reacting significantly with specific IgG antibodies, was selected. cDNA was subcloned and expressed in Escherichia coli. Sequencing of the cDNA revealed an open reading frame (ORF) of 1179 bases coding for a protein with an approximate molecular weight of 44 kDa. Immunoreactivity of the recombinant TS1 protein (rTS1) was evaluated by ELISA and Western blot analysis using pooled sera of ABPA patients. The rTS1 exhibited binding to specific IgG and IgE antibodies present in sera of ABPA patients. The deduced amino acid sequence showed homology to 60S ribosomal protein L3 (RpL3) of Aspergillus nidulans, Saccharomyces cerevisiae and Homo sapiens. The RpL3 of S. cerevesiae, tcm1, to which TS1 sequence shows significant homology (72% identity), is known to be responsible for conferring resistance against trichodermin (antibiotic, inhibiting protein synthesis). The present study has led to identification, cloning and expression of a 44-kDa novel allergen/antigen of Afu with sequence homology to L3 ribosomal protein with a probable role in resistance of Afu to antifungal drugs. Sixty-four per cent sequence identity of Afu RpL3 with human RpL3 and common regions in their predicted epitopes suggest a possibility of involvement of Afu RpL3 in autoimmune reactions due to molecular mimicry.
烟曲霉（Afu）是一种重要的真菌病原体，可引发过敏性和侵袭性呼吸系统疾病。Afu分泌的大量多功能变应原/抗原与发病机制有关。本研究采用cDNA文库方法来鉴定和表征新型Afu变应原/抗原。用过敏性支气管肺曲霉病（ABPA）患者的混合血清对Afu的cDNA文库进行免疫筛选。选择与特异性IgG抗体有显著反应的cDNA克隆TS1。将cDNA亚克隆并在大肠杆菌中表达。对该cDNA进行测序，发现一个1179个碱基的开放阅读框（ORF），编码一种分子量约为44 kDa的蛋白质。使用ABPA患者的混合血清，通过ELISA和蛋白质印迹分析评估重组TS1蛋白（rTS1）的免疫反应性。rTS1与ABPA患者血清中存在的特异性IgG和IgE抗体结合。推导的氨基酸序列与构巢曲霉、酿酒酵母和人类的60S核糖体蛋白L3（RpL3）具有同源性。酿酒酵母的RpL3即tcm1，TS1序列与其有显著同源性（72%同一性），已知其赋予对木霉菌素（一种抑制蛋白质合成的抗生素）的抗性。本研究已鉴定、克隆并表达了一种44 kDa的新型Afu变应原/抗原，其与L3核糖体蛋白具有序列同源性，可能在Afu对抗真菌药物的抗性中发挥作用。Afu RpL3与人类RpL3有64%的序列同一性，且在其预测表位中有共同区域，这表明由于分子模拟，Afu RpL3可能参与自身免疫反应。
