金耳和毛韧革菌麦角硫因生物合成基因的克隆及生物信 息学分析.

[Objective] To explore the biosynthetic pathway of ergothioneine in Naematelia aurantialba and Stereum hirsutum. [Method] The ergothioneine synthase gene Egt1 and Egt2 of N. aurantialba and S. hirsutum were cloned by PCR amplification technology, respectively, and their functions were analyzed by bioinformatics software. High performance liquid chromatography（HPLC）was used to identify the intermediate products of hercynine, ergothioneine and their contents in two species. [Result] The complete DNA sequences of Egt1 and Egt2 genes of two species were successfully cloned. The analysis using bioinformatics software revealed that Egt1 of the two species contained functional binding domains such as EgtD and SAM-dependent methyltransferase. Egt2 contained the binding sites for pyridoxal phosphate （LPL）and cysteine desulfurase. Further analysis indicated that Egt1 and Egt2 shared similar functional domains and substrate binding sites with model fungi（Schizosaccharomyces pombe and Neurospora crassa）. This result showed that Egt1 and Egt2 may have similar gene functions as these model fungi. HPLC analysis revealed the presence of hercynine and ergothioneine in N. aurantialba blastospore（JEYB）, S. hirsutum fermentum broth（ShFJY）, S. hirsutum mycelium（ShJST）and N. aurantialba fruiting bodies（JEZST）. Additionally, the ergothioneine content in the JEZST was found to be the highest at 113.19 μg/g, which was 7.45 times, 26.14 times, and 27.74 times higher than that of the JEYB, ShFJY, and ShJST, respectively. [Conclusion] The Egt1 and Egt2 genes of N. aurantialba and S. hirsutum were identified for the first time. It is hypothesized that the biosynthetic pathway of N. aurantialba and S. hirsutum are involved the catalysis of histidine by the Egt1 enzyme, resulting in the formation of hercynine. Subsequently, the Egt1 enzyme catalyzes the conversion of hercynine into hercynylcysteine sulfoxide. Finally, the Egt2 enzyme catalyzes the transformation of hercynylcysteine sulfoxide into ergothioneine. [ABSTRACT FROM AUTHOR]