scholarly journals Immunoelectron microscopic localization of the 210,000-mol wt microtubule-associated protein in cultured cells of primates.

1981 ◽  
Vol 91 (2) ◽  
pp. 438-445 ◽  
Author(s):  
M De Brabander ◽  
J C Bulinski ◽  
G Geuens ◽  
J De Mey ◽  
G G Borisy
Keyword(s):  
Molecular Weight ◽  
Intermediate Filaments ◽  
Sodium Azide ◽  
Cultured Cells ◽  
Entire Length ◽  
Ultrastructural Immunocytochemistry ◽  
Microscopic Localization ◽  
Protein Map ◽  
Tubulin Paracrystals

Results from ultrastructural immunocytochemistry on glutaraldehyde-fixed cells confirmed and extended findings previously obtained with immunofluorescence. A microtubule-associated protein (MAP) of 210,000 molecular weight was shown to be specifically associated with all cytoplasmic and mitotic microtubules along their entire length in primate cells. Specific labeling with the anti-MAP antibody could not be detected on any other subcellular structures, notably the centrosomes, kinetochores, microfilaments, and intermediate filaments. Treatment with the microtubule-disrupting drug, nocodazole, induced diffusion of the MAP throughout the cytoplasm. During repolymerization of microtubules following disassembly by nocodazole, the association of the MAP with the microtubules was intermediate and complete. When cells were treated with vinblastine, the tubulin paracrystals formed were heavily stained by the antibody. Neither sodium azide nor taxol affected the association of the MAP with microtubules.

Interactions between peripherin and neurofilaments in cultured cells: disruption of peripherin assembly by the NF-M and NF-H subunits

1999 ◽  
Vol 77 (1) ◽  
pp. 41-45 ◽  
Author(s):  
Jean-Martin Beaulieu ◽  
Janice Robertson ◽  
Jean-Pierre Julien
Keyword(s):  
Nervous System ◽  
Peripheral Nervous System ◽  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Cultured Cells ◽  
Intermediate Filament Protein ◽  
Physiological Conditions ◽  
Filament Protein ◽  
Filament Network ◽  
Filament Type

Neurofilaments are the principal intermediate filament type expressed by neurons. They are formed by the co-assembly of three subunits: NF-L, NF-M, and NF-H. Peripherin is another intermediate filament protein expressed mostly in neurons of the peripheral nervous system. In contrast to neurofilaments, peripherin can self-assemble to establish an intermediate filament network in cultured cells. The co-expression of neurofilaments and peripherin is found mainly during development and regeneration. We used SW13 cells devoid of endogenous cytoplasmic intermediate filaments to assess the exact assembly characteristics of peripherin with each neurofilament subunit. Our results demonstrate that peripherin can assemble with NF-L. In contrast, the co-expression of peripherin with the large neurofilament subunits interferes with peripherin assembly. These results confirm the existence of interactions between peripherin and neurofilaments in physiological conditions. Moreover, they suggest that perturbations in the stoichiometry of neurofilaments can have an impact on peripherin assembly in vivo.Key words: peripherin, neurofilament, SW13 cells, intermediate filament.

scholarly journals Contribution of quasi-fibrillar superstructures in peroxide quenching by collagen peptides derived from fish processing by-products and their application as natural food additives

2021 ◽  
Author(s):  
K Saleem ◽  
Pritha Dey ◽  
Charitha Sumeet ◽  
Mayur Bajaj ◽  
Y Geetika ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Cultured Cells ◽  
Food Additives ◽  
Lipid Peroxide ◽  
Natural Food ◽  
Type I ◽  
Collagen Hydrolysate ◽  
Simulated Digestion ◽  
Wide Range ◽  
Fibrillar Network

AbstractThis study attempts to identify the significant role played by the secondary structure of collagen-derived peptides that are involved in lipid peroxide quenching in food products. Collagen was extracted from the skin of Perch and swim bladder of Rohu at 45-78% efficiency. It was identified as type-I based on a high molecular weight (110kDa) and its ion-exchange elution profile. The collagen samples were enzymatically hydrolyzed and collagen hydrolysate (CH) was extracted with an efficiency of 0.67-0.74g/g of collagen. The CH samples displayed a molecular weight in the range of 8.2-9.7kDa and exhibited a higher abundance of charges resulting in higher solubility. The structural studies revealed that the CH peptides existed in polyproline-II helix and formed a mimic-triple helix in a wide range of pH. In neutral and alkaline pH, the mimic helices joined to form a hierarchical quasi-fibrillar network that was smaller than collagen fibrils but also more dynamic. The CH exhibited >95% degradation in 15h through simulated digestion. The CH were able to decrease peroxide formation by 84.5-98.9% in commercially available cod liver and almond oil and increased the shelf life of soya bean oil by a factor of 5 after 6 months of storage. The addition of CH to cultured cells quenched peroxide ions generated in situ and decreased stressor activity by a factor of 12. The reason behind the high efficacy of CH was deciphered to be the proximal charge stabilization by the quasi-fibrillar network, which allowed efficient peroxide quenching and long-term stability.

scholarly journals A cellular complex of BACE1 and γ-secretase sequentially generates Aβ from its full-length precursor

2019 ◽  
Vol 218 (2) ◽  
pp. 644-663 ◽  
Author(s):  
Lei Liu ◽  
Li Ding ◽  
Matteo Rovere ◽  
Michael S. Wolfe ◽  
Dennis J. Selkoe
Keyword(s):  
Molecular Weight ◽  
Cultured Cells ◽  
Amyloid Β ◽  
Full Length ◽  
Aβ Peptides ◽  
Whole Cells ◽  
Sequential Processing ◽  
Catalytically Active ◽  
Cellular Complex

Intramembrane proteolysis of transmembrane substrates by the presenilin–γ-secretase complex is preceded and regulated by shedding of the substrate’s ectodomain by α- or β-secretase. We asked whether β- and γ-secretases interact to mediate efficient sequential processing of APP, generating the amyloid β (Aβ) peptides that initiate Alzheimer’s disease. We describe a hitherto unrecognized multiprotease complex containing active β- and γ-secretases. BACE1 coimmunoprecipitated and cofractionated with γ-secretase in cultured cells and in mouse and human brain. An endogenous high molecular weight (HMW) complex (∼5 MD) containing β- and γ-secretases and holo-APP was catalytically active in vitro and generated a full array of Aβ peptides, with physiological Aβ42/40 ratios. The isolated complex responded properly to γ-secretase modulators. Alzheimer’s-causing mutations in presenilin altered the Aβ42/40 peptide ratio generated by the HMW β/γ-secretase complex indistinguishably from that observed in whole cells. Thus, Aβ is generated from holo-APP by a BACE1–γ-secretase complex that provides sequential, efficient RIP processing of full-length substrates to final products.

scholarly journals Assembly of type IV neuronal intermediate filaments in nonneuronal cells in the absence of preexisting cytoplasmic intermediate filaments

1993 ◽  
Vol 122 (6) ◽  
pp. 1323-1335 ◽  
Author(s):  
GY Ching ◽  
RK Liem
Keyword(s):  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Cultured Cells ◽  
Neuronal Cell ◽  
Intermediate Filament Protein ◽  
Amino Acid Residues ◽  
Intermediate Filament Proteins ◽  
Transfected Cells ◽  
Type Iv

We report here on the in vivo assembly of alpha-internexin, a type IV neuronal intermediate filament protein, in transfected cultured cells, comparing its assembly properties with those of the neurofilament triplet proteins (NF-L, NF-M, and NF-H). Like the neurofilament triplet proteins, alpha-internexin coassembles with vimentin into filaments. To study the assembly characteristics of these proteins in the absence of a preexisting filament network, transient transfection experiments were performed with a non-neuronal cell line lacking cytoplasmic intermediate filaments. The results showed that only alpha-internexin was able to self-assemble into extensive filamentous networks. In contrast, the neurofilament triplet proteins were incapable of homopolymeric assembly into filamentous arrays in vivo. NF-L coassembled with either NF-M or NF-H into filamentous structures in the transfected cells, but NF-M could not form filaments with NF-H. alpha-internexin could coassemble with each of the neurofilament triplet proteins in the transfected cells to form filaments. When all but 2 and 10 amino acid residues were removed from the tail domains of NF-L and NF-M, respectively, the resulting NF-L and NF-M deletion mutants retained the ability to coassemble with alpha-internexin into filamentous networks. These mutants were also capable of forming filaments with other wild-type neurofilament triplet protein subunits. These results suggest that the tail domains of NF-L and NF-M are dispensable for normal coassembly of each of these proteins with other type IV intermediate filament proteins to form filaments.

scholarly journals Intermediate filaments exchange subunits along their length and elongate by end-to-end annealing

2009 ◽  
Vol 185 (5) ◽  
pp. 769-777 ◽  
Author(s):  
Gülsen Çolakoğlu ◽  
Anthony Brown
Keyword(s):  
Intermediate Filaments ◽  
Actin Filaments ◽  
Fusion Proteins ◽  
Cultured Cells ◽  
Intermediate Filament Proteins ◽  
Subunit Exchange ◽  
End To End ◽  
Vimentin Intermediate Filament ◽  
Vimentin Filaments

Actin filaments and microtubules lengthen and shorten by addition and loss of subunits at their ends, but it is not known whether this is also true for intermediate filaments. In fact, several studies suggest that in vivo, intermediate filaments may lengthen by end-to-end annealing and that addition and loss of subunits is not confined to the filament ends. To test these hypotheses, we investigated the assembly dynamics of neurofilament and vimentin intermediate filament proteins in cultured cells using cell fusion, photobleaching, and photoactivation strategies in combination with conventional and photoactivatable fluorescent fusion proteins. We show that neurofilaments and vimentin filaments lengthen by end-to-end annealing of assembled filaments. We also show that neurofilaments and vimentin filaments incorporate subunits along their length by intercalation into the filament wall with no preferential addition of subunits to the filament ends, a process which we term intercalary subunit exchange.

scholarly journals New 1,2,3-Triazole Containing PolyestersviaClick Step-Growth Polymerization and Nanoparticles Made of Them

2018 ◽  
Vol 2018 ◽  
pp. 1-14 ◽  
Author(s):  
Tengiz Kantaria ◽  
Temur Kantaria ◽  
Giorgi Titvinidze ◽  
Giuli Otinashvili ◽  
Nino Kupatadze ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Sodium Azide ◽  
Dicarboxylic Acids ◽  
Solution Concentration ◽  
One Pot ◽  
Bromoacetic Acid ◽  
Aliphatic Polyesters ◽  
Step Growth ◽  
Step Growth Polymerization

High-molecular-weight AA-BB-type aliphatic polyesters were synthesizedviaCu(I)-catalyzed click step-growth polymerization (SGP) following a new synthetic strategy. The synthesis was performed between diyne and diazide monomers in an organic solvent as one pot process using three components and two stages. The dipropargyl esters of dicarboxylic acids (component 1) were used as diyne monomers, di-(bromoacetic acid)-alkylene diesters (component 2) were used as precursors of diazide monomers, and sodium azide (component 3) was used for generating diazide monomers. The SGP was carried out in two steps: at Step  1 dibromoacetates interacted with two moles of sodium azide resulting in diazide monomers which interacted in situ with diyne monomers at Step  2 in the presence of Cu(I) catalyst. A systematic study was done for optimizing the multiparameter click SGP in terms of the solvent, duration of both Step  1 and Step  2, solution concentration, catalyst concentration, catalyst and catalyst activator (ligand) nature, catalyst/ligand mole ratio, and temperature of both steps of the click SGP. As a result, high-molecular-weight (MWup to 74 kDa) elastic film-forming click polyesters were obtained. The new polymers were found suitable for fabricating biodegradable nanoparticles, which are promising as drug delivery containers in nanotherapy.

scholarly journals A Low-Molecular-Weight Antagonist for the Human Thyrotropin Receptor with Therapeutic Potential for Hyperthyroidism

Endocrinology ◽  
2008 ◽  
Vol 149 (12) ◽  
pp. 5945-5950 ◽  
Author(s):  
Susanne Neumann ◽  
Gunnar Kleinau ◽  
Stefano Costanzi ◽  
Susanna Moore ◽  
Jian-kang Jiang ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Cultured Cells ◽  
Therapeutic Potential ◽  
Partial Agonist ◽  
Primary Cultures ◽  
Graves Disease ◽  
Low Molecular Weight ◽  
Thyrotropin Receptor ◽  
Mrna Transcripts ◽  
Orally Active

Low-molecular-weight (LMW) antagonists for TSH receptor (TSHR) may have therapeutic potential as orally active drugs to block stimulating antibodies (TsAbs) in Graves’ hyperthyroidism. We describe an approach to identify LMW ligands for TSHR based on Org41841, a LMW partial agonist for the LH/choriogonadotropin receptor and TSHR. We used molecular modeling and functional experiments to guide the chemical modification of Org41841. We identified an antagonist (NIDDK/CEB-52) that selectively inhibits activation of TSHR by both TSH and TsAbs. Whereas initially characterized in cultured cells overexpressing TSHRs, the antagonist was also active under more physiologically relevant conditions in primary cultures of human thyrocytes expressing endogenous TSHRs in which it inhibited TSH- and TsAb-induced up-regulation of mRNA transcripts for thyroperoxidase. Our results establish this LMW compound as a lead for the development of higher potency antagonists and serve as proof of principle that LMW ligands that target TSHR could serve as drugs in patients with Graves’ disease.

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