Center for Biomedical Mass Spectrometry, Department of Biochemistry, Boston University School of Medicine, Boston, MA, USA; Laboratory of Insect Biochemistry, Department of Biochemistry, University of São Paulo, São Paulo, Brazil.
Center for Biomedical Mass Spectrometry, Department of Biochemistry, Boston University School of Medicine, Boston, MA, USA.
Comp Biochem Physiol B Biochem Mol Biol. 2020 Aug-Sep;246-247:110464. doi: 10.1016/j.cbpb.2020.110464. Epub 2020 Jun 14.
Spodoptera frugiperda is a widely distributed agricultural pest. It has previously been established that glycoproteins in the midgut microvillar membrane of insects are targets for toxins produced by different organisms as well as plant lectins. However, there is still little information about the N-glycome of membrane-bound midgut glycoproteins in Lepidoptera and other insect groups. The present study used mass spectrometry-based approaches to characterize the N-glycoproteins present in the midgut cell microvilli of Spodoptera frugiperda. We subjected midgut cell microvilli proteins to proteolytic digestion and enriched the resulting glycopeptides prior to analysis. We also performed endoglycosidase release of N-glycans in the presence of HO determining the compositions of released N-glycans by MALDI-TOF MS analysis and established the occupancy of the potential N-glycosylation sites. We report here a total of 160 glycopeptides, representing 25 N-glycan compositions associated with 70 sites on 35 glycoproteins. Glycan compositions consistent with oligomannose, paucimannose and complex/hybrid N-glycans represent 35, 30 and 35% of the observed glycans, respectively. The two most common N-glycan compositions were the complex/hybrid HexHexNAcdHex and the paucimannose structure that contains only the doubly-fucosylated trimannosylchitobiose core HexHexNAcdHex, each appearing in 22 occupied sites (13.8%). These findings enlighten aspects of the glycobiology of lepidopteran midgut microvilli.
斜纹夜蛾是一种广泛分布的农业害虫。先前已经证实，昆虫中肠微绒毛膜上的糖蛋白是不同生物体以及植物凝集素产生的毒素的靶标。然而，关于鳞翅目和其他昆虫群中膜结合中肠糖蛋白的 N-聚糖组，信息仍然很少。本研究使用基于质谱的方法来表征斜纹夜蛾中肠细胞微绒毛中的 N-糖蛋白。我们对中肠细胞微绒毛蛋白进行了蛋白水解消化，并在分析前对得到的糖肽进行了富集。我们还在 HO 存在的情况下进行了内切糖苷酶释放 N-聚糖，通过 MALDI-TOF MS 分析确定释放的 N-聚糖的组成，并确定潜在 N-糖基化位点的占有率。我们在这里报告了总共 160 个糖肽，代表与 35 种糖蛋白上的 70 个位点相关的 25 种 N-聚糖组成。与寡甘露糖、低聚糖和复杂/杂合 N-聚糖一致的聚糖组成分别占观察到的聚糖的 35%、30%和 35%。两种最常见的 N-聚糖组成是复杂/杂合的 HexHexNAcdHex 和仅包含双岩藻糖基化三甘露糖基核心 HexHexNAcdHex 的低聚糖结构，每个结构都出现在 22 个占据的位点（13.8%）。这些发现阐明了鳞翅目中肠微绒毛的糖生物学的一些方面。
