scholarly journals Site-specific analysis of N-glycans from different sheep prion strains

2021 ◽  
Vol 17 (2) ◽  
pp. e1009232
Author(s):  
Natali Nakić ◽  
Thanh Hoa Tran ◽  
Mislav Novokmet ◽  
Olivier Andreoletti ◽  
Gordan Lauc ◽  
...  
Keyword(s):  
Prion Protein ◽  
Prion Diseases ◽  
Mammalian Species ◽  
Site Specific ◽  
Prion Strains ◽  
Glycosylation Sites ◽  
Specific Analysis ◽  
Wide Range ◽  
The One ◽  
A Site

Prion diseases are a group of neurodegenerative diseases affecting a wide range of mammalian species, including humans. During the course of the disease, the abnormally folded scrapie prion protein (PrPSc) accumulates in the central nervous system where it causes neurodegeneration. In prion disorders, the diverse spectrum of illnesses exists because of the presence of different isoforms of PrPSc where they occupy distinct conformational states called strains. Strains are biochemically distinguished by a characteristic three-band immunoblot pattern, defined by differences in the occupancy of two glycosylation sites on the prion protein (PrP). Characterization of the exact N-glycan structures attached on either PrPC or PrPSc is lacking. Here we report the characterization and comparison of N-glycans from two different sheep prion strains. PrPSc from both strains was isolated from brain tissue and enzymatically digested with trypsin. By using liquid chromatography coupled to electrospray mass spectrometry, a site-specific analysis was performed. A total of 100 structures were detected on both glycosylation sites. The N-glycan profile was shown to be similar to the one on mouse PrP, however, with additional 40 structures reported. The results presented here show no major differences in glycan composition, suggesting that glycans may not be responsible for the differences in the two analyzed prion strains.

scholarly journals Bank vole prion protein extends the use of RT-QuIC assays to detect prions in a range of inherited prion diseases

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tze How Mok ◽  
Akin Nihat ◽  
Connie Luk ◽  
Danielle Sequeira ◽  
Mark Batchelor ◽  
...  
Keyword(s):  
Prion Protein ◽  
Bank Vole ◽  
Prion Diseases ◽  
Post Mortem ◽  
Prion Strains ◽  
Wide Range ◽  
Jakob Disease ◽  
Recombinant Prion Protein ◽  
First Time ◽  
Sporadic Cjd

AbstractThe cerebrospinal fluid (CSF) real-time quaking-induced conversion assay (RT-QuIC) is an ultrasensitive prion amyloid seeding assay for diagnosis of sporadic Creutzfeldt–Jakob disease (CJD) but several prion strains remain unexplored or resistant to conversion with commonly used recombinant prion protein (rPrP) substrates. Here, bank vole (BV) rPrP was used to study seeding by a wide range of archived post-mortem human CSF samples from cases of sporadic, acquired and various inherited prion diseases in high throughput 384-well format. BV rPrP substrate yielded positive reactions in 70/79 cases of sporadic CJD [Sensitivity 88.6% (95% CI 79.5–94.7%)], 1/2 variant CJD samples, and 9/20 samples from various inherited prion diseases; 5/57 non-prion disease control CSFs had positive reactions, yielding an overall specificity of 91.2% (95% CI 80.1–97.1%). Despite limitations of using post-mortem samples and our results’ discrepancy with other studies, we demonstrated for the first time that BV rPrP is susceptible to conversion by human CSF samples containing certain prion strains not previously responsive in conventional rPrPs, thus justifying further optimisation for wider diagnostic and prognostic use.

Site-Specific Ground Motion Studies for a Deep Soil Site Near Ahmedabad, Gujarat

2018 ◽  
pp. 31-65
Author(s):  
A. Boominathan ◽  
Krishna Kumar ◽  
R. Vijaya
Keyword(s):  
Seismic Hazard ◽  
Hazard Analysis ◽  
Response Analysis ◽  
Ground Response ◽  
Ground Response Analysis ◽  
Site Specific ◽  
Linear Approach ◽  
Specific Analysis ◽  
Soil Site ◽  
A Site

Design ground motions are usually developed by one of two approaches: by performing site-specific analyses or from provisions of building codes. Although contemporary codes consider the site effects to an extent, they provide more conservative estimates. Hence, site-specific analysis, which involves both the seismic hazard analysis and ground response analysis, is a preferred approach to obtain design ground motions. This chapter presents a site-specific analysis for a site near Ahmedabad, Gujarat. The seismic hazard analysis was carried out by DSHA approach. The site is predominantly characterized by deep stiff sandy clay deposits. Extensive shear wave velocity measurement is used for site classification and ground response analysis. The ground response analysis was carried out by using two approaches: the equivalent linear approach using SHAKE2000 and the non-linear approach using FLAC2D. The deep-stiff-soil site is found to amplify the ground motion. The response from nonlinear analysis is found to be considerably higher than those obtained from the equivalent linear approach.

scholarly journals A Site-Specific Analysis of the Implications of a Changing Ozone Profile and Climate for Stomatal Ozone Fluxes in Europe

2018 ◽  
Vol 230 (1) ◽  
Author(s):  
Felicity Hayes ◽  
Gina Mills ◽  
Rocio Alonso ◽  
Ignacio González-Fernández ◽  
Mhairi Coyle ◽  
...  
Keyword(s):  
Site Specific ◽  
Ozone Profile ◽  
Specific Analysis ◽  
Ozone Fluxes ◽  
A Site

scholarly journals Neuroinvasion in Prion Diseases: The Roles of Ascending Neural Infection and Blood Dissemination

2010 ◽  
Vol 2010 ◽  
pp. 1-16 ◽  
Author(s):  
Sílvia Sisó ◽  
Lorenzo González ◽  
Martin Jeffrey
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Prion Protein ◽  
Prion Diseases ◽  
Autonomic Nerves ◽  
Peripheral Organs ◽  
Prion Strains ◽  
New Variant ◽  
Jakob Disease ◽  
The Central Nervous System

Prion disorders are infectious, neurodegenerative diseases that affect humans and animals. Susceptibility to some prion diseases such as kuru or the new variant of Creutzfeldt-Jakob disease in humans and scrapie in sheep and goats is influenced by polymorphisms of the coding region of the prion protein gene, while other prion disorders such as fatal familial insomnia, familial Creutzfeldt-Jakob disease, or Gerstmann-Straussler-Scheinker disease in humans have an underlying inherited genetic basis. Several prion strains have been demonstrated experimentally in rodents and sheep. The progression and pathogenesis of disease is influenced by both genetic differences in the prion protein and prion strain. Some prion diseases only affect the central nervous system whereas others involve the peripheral organs prior to neuroinvasion. Many experiments undertaken in different species and using different prion strains have postulated common pathways of neuroinvasion. It is suggested that prions access the autonomic nerves innervating peripheral organs and tissues to finally reach the central nervous system. We review here published data supporting this view and additional data suggesting that neuroinvasion may concurrently or independently involve the blood vascular system.

scholarly journals The amino acid residue in position 163 of canine PrPC is critical to the exceptional resistance of dogs to prion infections: evidence from transgenic mouse models

2019 ◽  
Author(s):  
Enric Vidal ◽  
Natalia Fernández-Borges ◽  
Hasier Eraña ◽  
Beatriz Parra ◽  
Belén Pintado ◽  
...  
Keyword(s):  
Amino Acid ◽  
Mouse Model ◽  
Experimental Evidence ◽  
Transgenic Mouse ◽  
Prion Protein ◽  
Amino Acid Residue ◽  
Prion Diseases ◽  
Prion Infection ◽  
The One ◽  
Sheep Scrapie

ABSTRACTUnlike other species, such as cattle, cats or humans, prion disease has never been described in dogs, even though they were similarly exposed to the bovine spongiform encephalopathy (BSE) agent. This resistance prompted a thorough analysis of the canine PRNP gene and the presence of a negatively charged amino acid residue in position 163 was readily identified as potentially fundamental as it differed from all known susceptible species. Furthermore, recent results from our group demonstrated that mouse PRNP with the dog substitution N158D (mouse equivalent to position 163) rendered mice resistant to prion infection. In the present study, a transgenic mouse model was generated expressing dog prion protein (with glutamic acid at position 163) and challenged intracerebrally with a panel of prion isolates (including cattle BSE, sheep scrapie, atypical sheep scrapie, atypical BSE-L, sheep-BSE and chronic wasting disease, among others) none of which could infect them. The brains of these mice were subjected to in vitro prion amplification and failed to find even minimal amounts of misfolded prions providing definitive experimental evidence that dogs are resistant to prion disease. Subsequently, a second transgenic model was generated in which aspartic acid in position 163 was substituted for asparagine (the most common amino acid in this position in prion susceptible species) and this mutation resulted in susceptibility to BSE-derived isolates.These findings strongly support the hypothesis that the amino acid residue at position 163 of canine PrPC is a major determinant of the exceptional resistance of the canidae family to prion infection and establish this as a promising therapeutic target for prion diseases.AUTHOR SUMMARYCats, cattle, people and dogs were all exposed to mad cow disease but, unlike the other three, dogs never succumbed to the disease. We generated a mouse model expressing canine prion protein (instead of mouse prion protein) to provide experimental evidence that dogs are resistant to prion infection by challenging the mice with a panel of prion isolates. None of the prions could infect our transgenic mice that expressed dog prion protein. When the prion protein amino acid sequence of dogs was compared to that of other susceptible species, one amino acid in a specific position was found to be different to all the prion-susceptible animals. To determine if this amino acid was the one responsible for dogs’ resistance to prions, a second mouse model was generated with the canine prion protein but the critical amino acid was substituted for the one susceptible species have. When this model was challenged with the same panel of prions it could be infected with at least one of them. These results demonstrate the relevance of this amino acid position in determining susceptibility or resistance to prions, and this information can be used to design preventative treatments for prion diseases.

scholarly journals Ferritin Conjugates With Multiple Clickable Amino Acids Encoded by C-Terminal Engineered Pyrrolysyl-tRNA Synthetase

2021 ◽  
Vol 9 ◽  
Author(s):  
Yi-Hui Wang ◽  
Mu-Lung Jian ◽  
Pei-Jung Chen ◽  
Jo-Chu Tsou ◽  
Le P. Truong ◽  
...  
Keyword(s):  
Click Reaction ◽  
Trna Synthetase ◽  
Delivery Systems ◽  
Her2 Receptor ◽  
Site Specific ◽  
Protein Cage ◽  
Specific Loading ◽  
Wide Range ◽  
Multiple Drugs ◽  
A Site

This study reports the application of expanding genetic codes in developing protein cage-based delivery systems. The evolved Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS)•tRNAPyl pairs derived from directed evolution are examined to probe their recognition for para-substituted phenylalanine analogs. The evolved MmPylRS, AzFRS, harboring a wide range of substrates, is further engineered at the C-terminal region into another variant, AzFRS-MS. AzFRS-MS shows suppression of the elevated sfGFP protein amount up to 10 TAG stop codons when charging p-azido-l-phenylalanine (AzF, 4), which allows the occurrence of click chemistry. Since protein nanocages used as drug delivery systems that encompass multiple drugs through a site-specific loading approach remain largely unexplored, as a proof of concept, the application of AzFRS-MS for the site-specific incorporation of AzF on human heavy chain ferritin (Ftn) is developed. The Ftn-4 conjugate is shown to be able to load multiple fluorescence dyes or a therapeutic agent, doxorubicin (Dox), through the strain-promoted azide-alkyne cycloaddition (SPAAC) click reaction. Aiming to selectively target Her2+ breast cancer cells, Ftn-4-DOX conjugates fused with a HER2 receptor recognition peptide, anti-Her2/neu peptide (AHNP), is developed and demonstrated to be able to deliver Dox into the cell and to prolong the drug release. This work presents another application of evolved MmPylRS systems, whose potential in developing a variety of protein conjugates is noteworthy.

scholarly journals Genes encoding neuropeptide receptors are epigenetic markers in patients with head and neck cancer: a site-specific analysis

Oncotarget ◽  
2017 ◽  
Vol 8 (44) ◽  
pp. 76318-76328 ◽  
Author(s):  
Kiyoshi Misawa ◽  
Atsushi Imai ◽  
Daiki Mochizuki ◽  
Yuki Misawa ◽  
Shiori Endo ◽  
...  
Keyword(s):  
Head And Neck Cancer ◽  
Head And Neck ◽  
Neck Cancer ◽  
Site Specific ◽  
Epigenetic Markers ◽  
Neuropeptide Receptors ◽  
Specific Analysis ◽  
Genes Encoding ◽  
A Site

scholarly journals Site-specific glycosylation regulates the form and function of the intermediate filament cytoskeleton

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Heather J Tarbet ◽  
Lee Dolat ◽  
Timothy J Smith ◽  
Brett M Condon ◽  
E Timothy O'Brien ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Transferase Activity ◽  
Protein Protein Interactions ◽  
Site Specific ◽  
Form And Function ◽  
Glycosylation Sites ◽  
Wide Range ◽  
Dynamics And Function ◽  
Glcnac Transferase ◽  
And Function

Intermediate filaments (IF) are a major component of the metazoan cytoskeleton and are essential for normal cell morphology, motility, and signal transduction. Dysregulation of IFs causes a wide range of human diseases, including skin disorders, cardiomyopathies, lipodystrophy, and neuropathy. Despite this pathophysiological significance, how cells regulate IF structure, dynamics, and function remains poorly understood. Here, we show that site-specific modification of the prototypical IF protein vimentin with O-linked β-N-acetylglucosamine (O-GlcNAc) mediates its homotypic protein-protein interactions and is required in human cells for IF morphology and cell migration. In addition, we show that the intracellular pathogen Chlamydia trachomatis, which remodels the host IF cytoskeleton during infection, requires specific vimentin glycosylation sites and O-GlcNAc transferase activity to maintain its replicative niche. Our results provide new insight into the biochemical and cell biological functions of vimentin O-GlcNAcylation, and may have broad implications for our understanding of the regulation of IF proteins in general.

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