scholarly journals Site-specific N-glycosylation Characterization of Recombinant SARS-CoV-2 Spike Proteins

2020 ◽  
pp. mcp.RA120.002295 ◽  
Author(s):  
Yong Zhang ◽  
Wanjun Zhao ◽  
Yonghong Mao ◽  
Yaohui Chen ◽  
Shisheng Wang ◽  
...  
Keyword(s):  
Insect Cells ◽  
Host Cells ◽  
Protein Subunit ◽  
Genome Sequences ◽  
Complex Type ◽  
Site Specific ◽  
Differential Processing ◽  
S Protein ◽  
Shed Light

The glycoprotein spike (S) on the surface of SARS-CoV-2 is a determinant for viral invasion and host immune response. Herein, we characterized the site-specific N-glycosylation of S protein at the level of intact glycopeptides. All 22 potential N-glycosites were identified in the S-protein protomer and were found to be preserved among the 753 SARS-CoV-2 genome sequences. The glycosites exhibited glycoform heterogeneity as expected for a human cell-expressed protein subunit. We identified masses that correspond to 157 N-glycans, primarily of the complex type. In contrast, the insect cell-expressed S protein contained 38 N-glycans, completely of the high-mannose type. Our results revealed that the glycan types were highly determined by the differential processing of N-glycans among human and insect cells, regardless of the glycosites’ location. Moreover, the N-glycan compositions were conserved among different sizes of subunits. Our study indicate that the S protein N-glycosylation occurs regularly at each site, albeit the occupied N-glycans were diverse and heterogenous. This N-glycosylation landscape and the differential N-glycan patterns among distinct host cells are expected to shed light on the infection mechanism and present a positive view for the development of vaccines and targeted drugs.

scholarly journals Site-specific N-glycosylation Characterization of Recombinant SARS-CoV-2 Spike Proteins

2020 ◽  
Author(s):  
Yong Zhang ◽  
Wanjun Zhao ◽  
Yonghong Mao ◽  
Yaohui Chen ◽  
Shisheng Wang ◽  
...  
Keyword(s):  
Insect Cells ◽  
Host Cells ◽  
Genome Sequences ◽  
Complex Type ◽  
Site Specific ◽  
Differential Processing ◽  
S Protein ◽  
Protein Subunits ◽  
Shed Light

SummaryThe glycoprotein spike (S) on the surface of SARS-CoV-2 is a determinant for viral invasion and host immune response. Herein, we characterized the site-specific N-glycosylation of S protein at the level of intact glycopeptides. All 22 potential N-glycosites were identified in the S-protein protomer and were found to be preserved among the 753 SARS-CoV-2 genome sequences. The glycosites exhibited glycoform heterogeneity as expected for a human cell-expressed protein subunits. We identified masses that correspond to 157 N-glycans, primarily of the complex type. In contrast, the insect cell-expressed S protein contained 38 N-glycans, primarily of the high-mannose type. Our results revealed that the glycan types were highly determined by the differential processing of N-glycans among human and insect cells. This N-glycosylation landscape and the differential N-glycan patterns among distinct host cells are expected to shed light on the infection mechanism and present a positive view for the development of vaccines and targeted drugs.

scholarly journals Cryo-EM analysis of a feline coronavirus spike protein reveals a unique structure and camouflaging glycans

2020 ◽  
Vol 117 (3) ◽  
pp. 1438-1446 ◽  
Author(s):  
Tzu-Jing Yang ◽  
Yen-Chen Chang ◽  
Tzu-Ping Ko ◽  
Piotr Draczkowski ◽  
Yu-Chun Chien ◽  
...  
Keyword(s):  
Viral Entry ◽  
Protein Structures ◽  
Structural Features ◽  
Host Recognition ◽  
Complex Type ◽  
Site Specific ◽  
S Protein ◽  
Feline Infectious Peritonitis ◽  
Structural Insights ◽  
Feline Coronavirus

Feline infectious peritonitis virus (FIPV) is an alphacoronavirus that causes a nearly 100% mortality rate without effective treatment. Here we report a 3.3-Å cryoelectron microscopy (cryo-EM) structure of the serotype I FIPV spike (S) protein, which is responsible for host recognition and viral entry. Mass spectrometry provided site-specific compositions of densely distributed high-mannose and complex-type N-glycans that account for 1/4 of the total molecular mass; most of the N-glycans could be visualized by cryo-EM. Specifically, the N-glycans that wedge between 2 galectin-like domains within the S1 subunit of FIPV S protein result in a unique propeller-like conformation, underscoring the importance of glycosylation in maintaining protein structures. The cleavage site within the S2 subunit responsible for activation also showed distinct structural features and glycosylation. These structural insights provide a blueprint for a better molecular understanding of the pathogenesis of FIP.

Isolation and Characterization of a Two-Subunit Cytochromeb−c1Subcomplex fromRhodobacter capsulatusand Reconstitution of Its Ubihydroquinone Oxidation (Qo) Site with Purified Fe-S Protein Subunit†

Biochemistry ◽  
1998 ◽  
Vol 37 (46) ◽  
pp. 16242-16251 ◽  
Author(s):  
Maria B. Valkova-Valchanova ◽  
A. Sami Saribas ◽  
Brian R. Gibney ◽  
P. Leslie Dutton ◽  
Fevzi Daldal
Keyword(s):  
Protein Subunit ◽  
S Protein ◽  
Isolation And Characterization ◽  
Qo Site

Gelatin and Glycerine-Based Bioadhesive Vaginal Hydrogel

2020 ◽  
Vol 17 (4) ◽  
pp. 303-311
Author(s):  
Roberta Cassano ◽  
Federica Curcio ◽  
Delia Mandracchia ◽  
Adriana Trapani ◽  
Sonia Trombino
Keyword(s):  
Differential Scanning Calorimetry ◽  
Inflammatory Process ◽  
Vaginal Mucosa ◽  
Vaginal Infection ◽  
Scanning Calorimetry ◽  
Time Intervals ◽  
Site Specific ◽  
Release Studies ◽  
Hiv Seropositive

Aim: The work’s aim was the preparation and characterization of a hydrogel based on gelatin and glycerine, useful for site-specific release of benzydamine, an anti-inflammatory drug, able to attenuate the inflammatory process typical of the vaginal infection. Objective: The obtained hydrogel has been characterized by Electronic Scanning Microscopy (SEM) and Differential Scanning Calorimetry (DSC). In addition, due to the precursor properties, the hydrogel exhibits a relevant mucoadhesive activity. Methods: The swelling degree was evaluated at two different pHs and at defined time intervals. In particular, phosphate buffers were used at pH 6.6, in order to mimic the typical conditions of infectious diseases at the vaginal level, particularly for HIV-seropositive pregnant women, and pH 4.6, to simulate the physiological environment. Results: The obtained results revealed that the hydrogel swells up well at both pHs. Conclusion: Release studies conducted at both pathological and physiological pHs have shown that benzydamine is released at the level of the vaginal mucosa in a slow and gradual manner. These data support the hypothesis of the hydrogel use for the site-specific release of benzydamine in the vaginal mucosa.

scholarly journals Ni Oxidation State and Ligand Saturation Impact on the Capability of Octaazamacrocyclic Complexes to Bind and Reduce CO2

Molecules ◽  
2021 ◽  
Vol 26 (14) ◽  
pp. 4139
Author(s):  
Barbora Vénosová ◽  
Ingrid Jelemenská ◽  
Jozef Kožíšek ◽  
Peter Rapta ◽  
Michal Zalibera ◽  
...  
Keyword(s):  
Quantum Theory ◽  
Oxidation State ◽  
Central Atom ◽  
Population Analysis ◽  
Mulliken Population Analysis ◽  
Mulliken Population ◽  
Theoretical Methods ◽  
Ni Oxidation ◽  
Shed Light

Two 15-membered octaazamacrocyclic nickel(II) complexes are investigated by theoretical methods to shed light on their affinity forwards binding and reducing CO2. In the first complex 1[NiIIL]0, the octaazamacrocyclic ligand is grossly unsaturated (π-conjugated), while in the second 1[NiIILH]2+ one, the macrocycle is saturated with hydrogens. One and two-electron reductions are described using Mulliken population analysis, quantum theory of atoms in molecules, localized orbitals, and domain averaged fermi holes, including the characterization of the Ni-CCO2 bond and the oxidation state of the central Ni atom. It was found that in the [NiLH] complex, the central atom is reduced to Ni0 and/or NiI and is thus able to bind CO2 via a single σ bond. In addition, the two-electron reduced 3[NiL]2− species also shows an affinity forwards CO2.

scholarly journals Butyrophilin-like 2 regulates site-specific adaptations of intestinal γδ intraepithelial lymphocytes

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Casandra Panea ◽  
Ruoyu Zhang ◽  
Jeffrey VanValkenburgh ◽  
Min Ni ◽  
Christina Adler ◽  
...  
Keyword(s):  
Immune Responses ◽  
Intraepithelial Lymphocytes ◽  
Homeostatic Proliferation ◽  
Intestinal Epithelial ◽  
Intestinal Epithelial Barrier ◽  
Site Specific ◽  
Adaptive Immune ◽  
Intestinal Segments ◽  
Repertoire Diversity

AbstractTissue-resident γδ intraepithelial lymphocytes (IELs) orchestrate innate and adaptive immune responses to maintain intestinal epithelial barrier integrity. Epithelia-specific butyrophilin-like (Btnl) molecules induce perinatal development of distinct Vγ TCR+ IELs, however, the mechanisms that control γδ IEL maintenance within discrete intestinal segments are unclear. Here, we show that Btnl2 suppressed homeostatic proliferation of γδ IELs preferentially in the ileum. High throughput transcriptomic characterization of site-specific Btnl2-KO γδ IELs reveals that Btnl2 regulated the antimicrobial response module of ileal γδ IELs. Btnl2 deficiency shapes the TCR specificities and TCRγ/δ repertoire diversity of ileal γδ IELs. During DSS-induced colitis, Btnl2-KO mice exhibit increased inflammation and delayed mucosal repair in the colon. Collectively, these data suggest that Btnl2 fine-tunes γδ IEL frequencies and TCR specificities in response to site-specific homeostatic and inflammatory cues. Hence, Btnl-mediated targeting of γδ IEL development and maintenance may help dissect their immunological functions in intestinal diseases with segment-specific manifestations.

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