GfsA encodes a novel galactofuranosyltransferase involved in biosynthesis of galactofuranose antigen of O-glycan in Aspergillus nidulans and Aspergillus fumigatus.
GfsA encodes a novel galactofuranosyltransferase involved in biosynthesis of galactofuranose antigen of O-glycan in Aspergillus nidulans and Aspergillus fumigatus.
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DOI：
10.1111/mmi.12416
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发表时间：
2013-12
影响因子：
3.6
通讯作者：
Oka T
中科院分区：
文献类型：
--
作者：
Komachi Y;Hatakeyama S;Motomatsu H;Futagami T;Kizjakina K;Sobrado P;Ekino K;Takegawa K;Goto M;Nomura Y;Oka T
曲霉属丝状真菌的细胞壁含有含呋喃半乳糖的多糖和糖复合物，包括 O-聚糖、N-聚糖、真菌型半乳甘露聚糖和糖基肌醇磷酸神经酰胺，它们对细胞壁的完整性很重要。在这里，我们试图鉴定将呋喃半乳糖单体偶联到构巢曲霉中其他壁成分上的呋喃半乳糖基转移酶。使用反向遗传和生化方法，我们确定 AN8677 基因编码呋喃半乳糖基转移酶，我们将其称为 GfsA，参与呋喃半乳糖 (Galf) 抗原的生物合成。 gfsA 的破坏减少了 β-Galf 特异性抗体 EB-A2 与 O-糖基化 WscA 蛋白和半乳甘露糖蛋白的结合。体外呋喃半乳糖抗原合酶测定结果表明，GfsA 对糖蛋白中的 O-聚糖具有 β1,5- 或 β1,6- 呋喃半乳糖基转移酶活性，使用 UDP-D-呋喃半乳糖作为糖供体，并需要二价锰阳离子来发挥活性。根据细胞分级实验，发现 GfsA 定位于高尔基体。 ΔgfsA 细胞表现出异常形态，其特征是菌丝延伸不良、菌丝弯曲和分生孢子形成有限。在子囊菌门的 Pezizomycotina 亚门成员中鉴定出了几种 gfsA 直系同源物，包括人类病原体烟曲霉。据我们所知，这是真菌β-呋喃半乳糖基转移酶的首次表征，该酶被证明参与高尔基体中O-聚糖的呋喃半乳糖抗原生物合成。
The cell walls of filamentous fungi in the genus Aspergillus have galactofuranose-containing polysaccharides and glycoconjugates, including O-glycans, N-glycans, fungal-type galactomannan, and glycosylinositolphosphoceramide, which are important for cell wall integrity. Here, we attempted to identify galactofuranosyltransferases that couple galactofuranose monomers onto other wall components in Aspergillus nidulans. Using reverse-genetic and biochemical approaches, we identified that the AN8677 gene encoded a galactofuranosyltransferase, which we called GfsA, involved in galactofuranose (Galf) antigen biosynthesis. Disruption of gfsA reduced binding of β-Galf-specific antibody EB-A2 to O-glycosylated WscA protein and galactomannoproteins. The results of an in-vitro galactofuranose antigen synthase assay revealed that GfsA has β1,5- or β1,6- galactofuranosyltransferase activity for O-glycans in glycoproteins, uses UDP-D-galactofuranose as a sugar donor, and requires a divalent manganese cation for activity. GfsA was found to be localized at the Golgi apparatus based on cellular fractionation experiments. ΔgfsA cells exhibited an abnormal morphology characterized by poor hyphal extension, hyphal curvature, and limited formation of conidia. Several gfsA orthologs were identified in members of the Pezizomycotina subphylum of Ascomycota, including the human pathogen Aspergillus fumigatus. To our knowledge, this is the first characterization of a fungal β-galactofuranosyltransferase, which was shown to be involved in galactofuranose antigen biosynthesis of O-glycans in the Golgi.