LitCovid-sample-sentences  

PMC:7443692 / 13791-24021 JSONTXT 12 Projects

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Id Subject Object Predicate Lexical cue
T74 0-105 Sentence denotes Glycomics-Informed Glycoproteomics Reveals Site-Specific Microheterogeneity of SARS-CoV-2 S Glycosylation
T75 106-190 Sentence denotes We utilized multiple approaches to examine glycosylation of the SARS-CoV-2 S trimer.
T76 191-326 Sentence denotes First, the portfolio of glycans linked to SARS-CoV-2 S trimer immunogen was analyzed after their release from the polypeptide backbone.
T77 327-453 Sentence denotes N-glycans were released from protein by treatment with PNGase F- and O-glycans were subsequently released by beta-elimination.
T78 454-650 Sentence denotes After permethylation to enhance detection sensitivity and structural characterization, released glycans were analyzed by multi-stage mass spectrometry (MSn) (Aoki et al., 2007; Aoki et al., 2008).
T79 651-776 Sentence denotes Mass spectra were processed by GRITS Toolbox, and the resulting annotations were validated manually (Weatherly et al., 2019).
T80 777-933 Sentence denotes Glycan assignments were grouped by type and by additional structural features for relative quantification of profile characteristics (Figure 2 A; Table S3).
T81 934-1102 Sentence denotes This analysis quantified 49 N-glycans and revealed that 55% of the total glycan abundance was of the complex type, 17% was of the hybrid type, and 28% was high mannose.
T82 1103-1252 Sentence denotes Among the complex and hybrid N-glycans, we observed a high degree of core fucosylation and significant abundance of bisected and LacDiNAc structures.
T83 1253-1494 Sentence denotes We also observed sulfated N-linked glycans by using negative mode MSn analyses (Table S13), although signal intensity was too low in positive ion mode (at least 10-fold lower than any of the non-sulfated glycans) for accurate quantification.
T84 1495-1582 Sentence denotes In addition, we detected 15 O-glycans released from the S trimer (Figure S5; Table S4).
T85 1583-1721 Sentence denotes Figure 2 Glycomics-Informed Glycoproteomics Reveals Substantial Site-Specific Microheterogeneity of N-linked Glycosylation on SARS-CoV-2 S
T86 1722-1825 Sentence denotes (A) Glycans released from SARS-CoV-2 S protein trimer immunogen were permethylated and analyzed by MSn.
T87 1826-1947 Sentence denotes Structures were assigned and grouped by type and structural features, and prevalence was determined based on ion current.
T88 1948-2006 Sentence denotes The pie chart shows basic division by broad N-glycan type.
T89 2007-2075 Sentence denotes The bar graph provides additional detail about the glycans detected.
T90 2076-2249 Sentence denotes The most abundant structure with a unique categorization by glycomics for each N-glycan type in the pie chart, or above each feature category in the bar graph, is indicated.
T91 2250-2474 Sentence denotes (B–E) Glycopeptides were prepared from SARS-CoV-2 S protein trimer immunogen by using multiple combinations of proteases, analyzed by LC-MSn, and the resulting data were searched by using several different software packages.
T92 2475-2597 Sentence denotes Four representative sites of N-linked glycosylation with specific features of interest were chosen and are presented here.
T93 2598-2826 Sentence denotes N0074 (B) and N0149 (C) are shown that occur in variable insert regions of S compared to SARS-CoV and other related coronaviruses, and there are emerging variants of SARS-CoV-2 that disrupt these two sites of glycosylation in S.
T94 2827-2885 Sentence denotes N0234 (D) contains the most high-mannose N-linked glycans.
T95 2886-3036 Sentence denotes N0801 (D) is an example of glycosylation in the S2 region of the immunogen and displays a high degree of hybrid glycosylation compared to other sites.
T96 3037-3119 Sentence denotes The abundance of each composition is graphed in terms of assigned spectral counts.
T97 3120-3240 Sentence denotes Representative glycans (as determined by glycomics analysis) for several abundant compositions are shown in SNFG format.
T98 3241-3372 Sentence denotes The abbreviations used here and throughout the manuscript are as follows: N, HexNAc; H, hexose; F, fucose; A, Neu5Ac; S, sulfation.
T99 3373-3537 Sentence denotes Note that the graphs for the other 18 sites and other graphs grouping the microheterogeneity observed by other properties are presented in Supplemental Information.
T100 3538-3778 Sentence denotes To determine occupancy of N-linked glycans at each site, we employed a sequential deglycoslyation approach by using Endoglycosidase H and PNGase F in the presence of 18O-H2O after tryptic digestion of S (Wang et al., 2020; Yu et al., 2018).
T101 3779-3910 Sentence denotes After LC-MS/MS analyses, the resulting data confirmed that 19 of the canonical sequons had occupancies greater than 95% (Table S5).
T102 3911-4075 Sentence denotes One canonical sequence, N0149, had insufficient spectral counts for quantification by this method, but subsequent analyses described below suggested high occupancy.
T103 4076-4181 Sentence denotes The two most C-terminal N-linked sites, N1173 and N1194, had reduced occupancy, 52% and 82% respectively.
T104 4182-4361 Sentence denotes Reduced occupancy at these sites could reflect hindered en bloc transfer by the oligosaccharyltransferase (OST) due to primary amino acid sequences at or near the N-linked sequon.
T105 4362-4694 Sentence denotes Alternatively, this could reflect these two sites being post-translationally modified after release of the protein by the ribosome by a less efficient STT3B-containing OST, either due to activity or initial folding of the polypeptide, as opposed to co-translationally modified by the STT3A-containing OST (Ruiz-Canada et al., 2009).
T106 4695-5021 Sentence denotes None of the non-canonical sequons (three N-X-C sites and four N-G-L/I/V sites; Zielinska et al., 2010) showed significant occupancy (>5%), except for N0501, which showed moderate (19%) conversion to 18O-Asp that could be due to deamidation that is facilitated by glycine at the +1 position (Table S5) (Palmisano et al., 2012).
T107 5022-5177 Sentence denotes Further analysis of this site (see below) by direct glycopeptide analyses allowed us to determine that N0501 undergoes deamidation but is not glycosylated.
T108 5178-5339 Sentence denotes Thus, all, and only the, 22 canonical sequences for N-linked glycosylation (N-X-S/T) are utilized, with only N1173 and N1194 demonstrating occupancies below 95%.
T109 5340-5502 Sentence denotes Next, we applied three different proteolytic digestion strategies to the SARS-CoV-2 S immunogen to maximize glycopeptide coverage by subsequent LC-MS/MS analyses.
T110 5503-5803 Sentence denotes Extended gradient nanoflow reverse-phase LC-MS/MS was carried out on a ThermoFisher Lumos Tribrid instrument using Step-HCD fragmentation on each of the samples (see STAR Methods for details, as well as Duan et al., 2018; Escolano et al., 2019; Wang et al., 2020; Yu et al., 2018; Zhou et al., 2017).
T111 5804-6117 Sentence denotes After data analyses using pGlyco 2.2.2 (Liu et al., 2017), Byonic (Bern et al., 2012), and manual validation of glycan compositions against our released glycomics findings (Figure 2A; Tables S3 and S13), we were able to determine the microheterogeneity at each of the 22 canonical sites (Figures 2B–2E; Table S6).
T112 6118-6226 Sentence denotes Notably, none of the non-canonical consensus sequences, including N0501, displayed any quantifiable glycans.
T113 6227-6354 Sentence denotes The N-glycosites N0074 (Figure 2B) and N0149 (Figure 2C) are highly processed and display a typical mammalian N-glycan profile.
T114 6355-6445 Sentence denotes N0149 is, however, modified with several hybrid N-glycan structures, whereas N0074 is not.
T115 6446-6636 Sentence denotes N0234 (Figure 2D) and N0801 (Figure 2E) have N-glycan profiles more similar to those found on other viruses such as HIV (Watanabe et al., 2019) that are dominated by high-mannose structures.
T116 6637-6791 Sentence denotes N0234 (Figure 2D) displays an abundance of Man7-Man9 high-mannose structures, suggesting stalled processing by early-acting ER and cis-Golgi mannosidases.
T117 6792-6989 Sentence denotes In contrast, N0801 (Figure 2E) is processed more efficiently to Man5 high-mannose and hybrid structures, suggesting that access to the glycan at this site by MGAT1 and α-Mannosidase II is hindered.
T118 6990-7257 Sentence denotes In general, for all 22 sites (Figures 2B–2E; Table S6), we observed underprocessing of complex glycan antennae (i.e., under-galactosylation and under-sialylation) and a high degree of core fucosylation in agreement with released glycan analyses (Figure 2A; Table S3).
T119 7258-7356 Sentence denotes We also observed a small percent of sulfated N-linked glycans at several sites (Tables S6 and S8).
T120 7357-7572 Sentence denotes Based on the assignments and the spectral counts for each topology, we were able to determine the percent of total N-linked glycan types (high-mannose, hybrid, or complex) present at each site (Figure 3 ; Table S7).
T121 7573-7819 Sentence denotes Notably, three of the sites (N0234, N0709, and N0717) displayed more than 50% high-mannose glycans, whereas 11 other sites (N0017, N0074, N0149, N0165, N0282, N0331, N0657, N1134, N1158, N1173, and N1194) were more than 90% complex when occupied.
T122 7820-7886 Sentence denotes The other eight sites were distributed between these two extremes.
T123 7887-8017 Sentence denotes Notably, only one site (N0717 at 45%), which also had greater than 50% high mannose (55%), had greater than 33% hybrid structures.
T124 8018-8288 Sentence denotes To further evaluate the heterogeneity, we grouped all the topologies into the 20 classes recently described by the Crispin laboratory, adding two categories (sulfated and unoccupied) that we refer to here as the Oxford classification (Table S8) (Watanabe et al., 2020a).
T125 8289-8588 Sentence denotes Among other features observed, this classification allowed us to observe that although most sites with high-mannose structures were dominated by the Man5GlcNAc2 structure, N0234 and N0717 were dominated by the higher Man structures of Man8GlcNAc2 and Man7GlcNAc2, respectively (Figure S7; Table S8).
T126 8589-8812 Sentence denotes Limited processing at N0234 is in agreement with a recent report suggesting that high-mannose structures at this site help to stabilize the receptor-binding domain of S (www.biorxiv.org/content/10.1101/2020.06.11.146522v1).
T127 8813-9140 Sentence denotes Furthermore, applying the Oxford classifications to our dataset clearly demonstrates that the three most C-terminal sites (N1158, N1173, and N1194), dominated by complex-type glycans, were more often further processed (i.e., multiple antennae) and elaborated (i.e., galactosylation and sialylation) than other sites (Table S8).
T128 9141-9235 Sentence denotes Figure 3 SARS-CoV-2 S Immunogen N-glycan Sites Are Predominantly Modified by Complex N-glycans
T129 9236-9517 Sentence denotes N-glycan topologies were assigned to all 22 sites of the S protomer and the spectral counts for each of the three types of N-glycans (high-mannose, hybrid, and complex), as well as the unoccupied peptide spectral match counts at each site, were summed and visualized as pie charts.
T130 9518-9605 Sentence denotes Note that only N1173 and N1194 show an appreciable amount of the unoccupied amino acid.
T131 9606-9889 Sentence denotes We also analyzed our generated mass spectrometry data for the presence of O-linked glycans based on our glycomic findings (Figure S5; Table S4) and a recent manuscript suggesting significant levels of O-glycosylation of S1 and S2 when expressed independently (Shajahan et al., 2020).
T132 9890-10026 Sentence denotes We were able to confirm sites of O-glycan modification with microheterogeneity observed for the vast majority of these sites (Table S9).
T133 10027-10230 Sentence denotes However, occupancy at each site, determined by spectral counts, was observed to be very low (below 4%), except for Thr0323, which had a modestly higher but still low 11% occupancy (Figure S6; Table S10).