scholarly journals Microalgal protein AstaP is a potent carotenoid solubilizer and delivery module with a broad carotenoid binding repertoire

2021 ◽  
Author(s):  
Yury B Slonimskiy ◽  
Nikita A Egorkin ◽  
Thomas N. Friedrich ◽  
Eugene G. Maksimov ◽  
Nikolai N. Sluchanko
Keyword(s):  
Escherichia Coli ◽  
Neurodegenerative Disorders ◽  
Biomedical Applications ◽  
Protein Complexes ◽  
Water Soluble ◽  
Biological Functions ◽  
Cost Efficient ◽  
Delivery Module ◽  
Β Carotene

Carotenoids are lipophilic substances with many biological functions, from coloration to photoprotection. Being potent antioxidants, carotenoids have multiple biomedical applications, including the treatment of neurodegenerative disorders and retina degeneration. Nevertheless, the delivery of carotenoids is substantially limited by their poor solubility in the aqueous phase. Natural water-soluble carotenoproteins can facilitate this task, necessitating studies on their ability to uptake and deliver carotenoids. One such promising carotenoprotein, AstaP (Astaxanthin-binding protein), was recently identified in eukaryotic microalgae, but its structure and functional properties remained largely uncharacterized. By using a correctly folded recombinant protein, here we show that AstaP is an efficient carotenoid solubilizer that can stably bind not only astaxanthin but also zeaxanthin, canthaxanthin, and, to a lesser extent, β-carotene, i.e. carotenoids especially valuable to human health. AstaP accepts carotenoids provided as acetone solutions or embedded in membranes, forming carotenoid-protein complexes with an apparent stoichiometry of 1:1. We successfully produced AstaP holoproteins in specific carotenoid-producing strains of Escherichia coli, proving it is amenable to cost-efficient biotechnology processes. Regardless of the carotenoid type, AstaP remains monomeric in both apo- and holoforms, while its rather minimalistic mass (~20 kDa) makes it an especially attractive antioxidant delivery module. In vitro, AstaP transfers different carotenoids to the liposomes and to unrelated proteins from cyanobacteria, which can modulate their photoactivity and/or oligomerization. These findings expand the toolkit of the characterized carotenoid-binding proteins and outline the perspective of the use of AstaP as a unique monomeric antioxidant nanocarrier with an extensive carotenoid-binding repertoire.

Exploring the Pharmacognostic Features, Anti-oxidant and Lipid Lowering Potential of Fagopyrum esculentum Moench. Seed

2020 ◽  
Vol 6 (2) ◽  
pp. 155-169
Author(s):  
Neeraj Panihar ◽  
Neeru Vasudeva ◽  
Sunil Sharma ◽  
Babu Lal Jangir
Keyword(s):  
Ethyl Acetate ◽  
Wistar Rats ◽  
Ethanol Extract ◽  
Water Soluble ◽  
Fagopyrum Esculentum ◽  
Lipid Lowering ◽  
Standard Drug ◽  
Fagopyrum Esculentum Moench ◽  
Β Carotene

Background: Fagopyrum esculentum Moench. is a herb consumed as food and has medicinal value. It is a rich source of bioactive nutrients which cure and prevent many ailments. Traditionally, it is used to treat hypertension, diabetes, constipation, cancer etc. Methods and Objective: Present work illustrates morphological, microscopic and physicochemical parameters of Fagopyrum esculentum seeds as per WHO guidelines, in vitro antioxidant activity; assessed by DPPH scavenging method, hydrogen peroxide scavenging assay and β-carotene linoleic acid bleaching method and study of lipid lowering potential of the ethyl acetate and ethanol extract of seeds on normal diet fed Wistar rats. Results: Morphological studies delineated the triangular shape, dark brown colour, 8 mm length and 6 mm width of the seed. The microscopic examination of the transverse section of seed depicted features like testa or pericarp (seed coat), the endosperm, embryo and sclerenchyma cells. Study of physiochemical parameters exhibited 0.3±0.02% of foreign matter and 1.44±0.51% crude fibre content. Total ash, acid insoluble ash and water soluble ash value were 6.7±1.7%, 1.9±0.23% and 3.9± 0.31% respectively. Alcohol soluble and water soluble extractive value came out to be 65.02± 3.21 mg/g and 12.7±1.24 mg/g respectively. Foaming index was less than 100, swelling index was found to be 0.5±0.01 ml/g. Loss on drying was 4.02±1.27%. Phytochemical screening of ethyl acetate and ethanol extract revealed the presence of alkaloids, carbohydrates, phenolic compounds, phytosterols and flavonoids. Trace amount of heavy metals (arsenic, cadmium, lead, mercury) were determined by atomic absorption spectrophotometer. Pesticide residue analysis confirmed the presence of nontoxic pesticides like dimethipin, hymexazol, phenothrin-2, methoprene, triadimenol, prohydrojasmon- 1, jasmolin ii, triademinol, jasmolin i, prohydrojasmone i, cyromazine in both the extracts by gc-ms spectrometer. The ethyl acetate and ethanol extract has shown significant in-vitro antioxidant activities demonstrated by the DPPH method (IC50 = 94.37±2.51 and 216.04±4.39 μg/ml respectively), hydrogen peroxide scavenging assay (IC50 = 83.72±3.72 and 193.47±5.05 µg/ml respectively) and β-carotene bleaching method (IC50 = 100.67±4.01 and 205.39±2.89 µg/ml respectively). Lipid lowering study performed on Wistar rats demonstrated a significant (p<0.001) decrease in serum Total Cholesterol (TC), Triglyceride (TG) and increase in High Density Lipoprotein (HDL) level as compared to normal group. Both the extracts have shown a non significant difference in the level of TG as compared to standard drug atorvastatin, depicting that the efficacy of extracts is at par with that of standard drug atorvastatin. Conclusion: Pharmacognostical study of the plant can be a very good tool for identification as well as authentication of a herb. Moreover, these parameters may be helpful in the development of monograph of the plant. Pharmacological activity confirmed Fagopyrum esculentum Moench. seed to be a good antioxidant and have lipid lowering potential.

scholarly journals Water Soluble Pleurotus ostreatus Polysaccharide Down-Regulates the Expression of MMP-2 and MMP-9 in Caco-2 Cells

2013 ◽  
Vol 41 (2) ◽  
pp. 553
Author(s):  
Sabina COJOCARU ◽  
Mihaela RADU ◽  
Liviu Gabriel BODEA ◽  
Mirela Mihaela CIMPEANU ◽  
Gogu GHEORGHITA ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Cancer Cells ◽  
Protein Complexes ◽  
Gelatin Zymography ◽  
Water Soluble ◽  
Colon Cancer Cells ◽  
Sod Activity ◽  
Kinetic Measurements ◽  
The Third

Many polysaccharides and polysaccharide-protein complexes isolated from mushrooms have immunomodulatory and anti-cancer effects. Our aim was to study the regulatory mechanisms of Caco-2 cell response to water soluble P. ostreatus polysaccharide extract up to 72 hours. Specific enzymatic activities were assessed by kinetic measurements. The reduced glutathione content and the lipid peroxidation level were also analyzed. Protein expression of several heat shock proteins, Bcl-2 and metalloproteinases 2 and 9 were revealed by Western blot. Gelatin zymography assay was used to evaluate the MMP-2 and MMP-9 activities. Until the third day of exposure the total SOD activity decreased continuously by 30%, whereas GST and GR ones diminished by 17% respectively 30.5% compared to control. No significant changes were observed in CAT and G6PDH specific activities as well as in GSH and MDA concentration. After the third day of exposure a significant up-regulation of Hsp60 and Hsp90 expression and a down-regulation of Hsp70 one were registered. Bcl-2 protein levels were down-regulated by 50% in the first day of treatment but increased after 3 days. MMP-2 and 9 secretion in the culture medium was significantly reduced suggesting a diminished ability of invasion of colon cancer cells. Our data revealed that in vitro treatment with P. ostreatus aqueous polysaccharide extract does not induce apoptosis in Caco-2 cell line but it could inhibit the invasion of colon cancer cells through the basement membrane.

Purification of complexes of nuclear oncogene p53 with rat and Escherichia coli heat shock proteins: in vitro dissociation of hsc70 and dnaK from murine p53 by ATP

1988 ◽  
Vol 8 (3) ◽  
pp. 1206-1215
Author(s):  
C F Clarke ◽  
K Cheng ◽  
A B Frey ◽  
R Stein ◽  
P W Hinds ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Protein Interactions ◽  
Protein Complexes ◽  
P53 Protein ◽  
Immunoaffinity Column ◽  
E Coli ◽  
Shock Proteins

Oligomeric protein complexes containing the nuclear oncogene p53 and the simian virus 40 large tumor antigen (D. I. H. Linzer and A. J. Levine, Cell 17:43-51, 1979), the adenovirus E1B 55-kilodalton (kDa) tumor antigen, and the heat shock protein hsc70 (P. Hinds, C. Finlay, A. Frey, and A. J. Levine, Mol. Cell. Biol. 7:2863-2869, 1987) have all been previously described. To begin isolating, purifying, and testing these complexes for functional activities, we have developed a rapid immunoaffinity column purification. p53-protein complexes are eluted from the immunoaffinity column by using a molar excess of a peptide comprising the epitope recognized by the p53 monoclonal antibody. This mild and specific elution condition allows p53-protein interactions to be maintained. The hsc70-p53 complex from rat cells is heterogeneous in size, with some forms of this complex associated with a 110-kDa protein. The maximum apparent molecular mass of such complexes is 660,000 daltons. Incubation with micromolar levels of ATP dissociates this complex in vitro into p53 and hsc70 110-kDa components. Nonhydrolyzable substrates of ATP fail to promote this dissociation of the complex. Murine p53 synthesized in Escherichia coli has been purified 660-fold on the same antibody affinity column and was found to be associated with an E. coli protein of 70 kDa. Immunoblot analysis with specific antisera demonstrated that this E. coli protein was the heat shock protein dnaK, which has extensive sequence homology with the rat hsc70 protein. Incubation of the immunopurified p53-dnaK complex with ATP resulted in the dissociation of the p53-dnaK complex as it did with the p53-hsc70 complex. This remarkable conservation of p53-heat shock protein interactions and the specificity of dissociation reactions suggest a functionally important role for heat shock proteins in their interactions with oncogene proteins.

scholarly journals Composite Scaffolds Based on Silver Nanoparticles for Biomedical Applications

2015 ◽  
Vol 2015 ◽  
pp. 1-8 ◽  
Author(s):  
Jenel Marian Patrascu ◽  
Ioan Avram Nedelcu ◽  
Maria Sonmez ◽  
Denisa Ficai ◽  
Anton Ficai ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Silver Nanoparticles ◽  
Composite Materials ◽  
Chemical Reaction ◽  
Biomedical Applications ◽  
Chemical Reduction ◽  
Composite Scaffolds ◽  
Sputtering Deposition ◽  
Plasma Sputtering

This paper presents the synthesis, characterisation, andin vitrotesting of homogenous and heterogeneous materials containing silver nanoparticles (nanoAg). Three types of antiseptic materials based on collagen (COLL), hydroxyapatite (HA), and collagen/hydroxyapatite (COLL/HA) composite materials were obtained. The synthesis of silver nanoparticles was realized by chemical reaction as well as plasma sputtering deposition. The use of chemical reduction allows the synthesis of homogenous materials while the plasma sputtering deposition can be easily used for the synthesis of homogeneous and heterogeneous support. Based on thein vitroassays clear antiseptic activity againstEscherichia coliwas relieved even at low content of nanoAg (10 ppm).

Mitomycin C-Activity Effected by Vitamins B1, C, E and β-Carotene under Irradiation with γ-Rays

2003 ◽  
Vol 58 (3-4) ◽  
pp. 244-248 ◽  
Author(s):  
Edith Heinrich ◽  
Nikola Getoff
Keyword(s):  
Escherichia Coli ◽  
Mitomycin C ◽  
Vitamin B1 ◽  
Fold Increase ◽  
Oh Radicals ◽  
Strong Decrease ◽  
Γ Rays ◽  
Escherichia Coli Bacteria ◽  
Β Carotene

Vitamin B1 (thiamine) can essentially effect the activity of mitomycin C (MMC), added individually or in combination with antioxidant vitamins (C, E-acetate, β-carotene) as found in experiments in vitro (Escherichia coli bacteria, AB 1157) under irradiation with γ-rays. The environment plays a crucial role. In airfree media vitamin B1 leads to a 2-fold increase of the MMC-efficiency, but adding vitamin C it decreases. In the presence of all vitamins (B1, C, E-ac., and β-carotene) the MMC-action increases about 1.8-fold. In aerated media vitamin B1 causes an about 4-times increase of the MMC-efficiency, but by adding vitamin B1 and C the MMC-activity decreases by a factor of two, whereas in the presence of B1, C, E-ac., and β-carotene it rises again to 2.6-fold. In environment saturated with N2O (conversion of e-aq into OH radicals) a different picture is observed. The presence of vitamin B1 or vitamin B1 + C causes a strong decrease of the MMC-efficiency, but the addition of all vitamins (B1, C, E-ac., and β-car.) leads to a small increase of the cytostatic action. The results demonstrate the influence of vitamin B1 used individually or in combination with other antioxidants on the MMC-efficiency and the strong effect of the environment. The results are of interest for the application of MMC in radiotherapy.

scholarly journals Tol-Dependent Macromolecule Import through the Escherichia coli Cell Envelope Requires the Presence of an Exposed TolA Binding Motif

2005 ◽  
Vol 187 (21) ◽  
pp. 7526-7534 ◽  
Author(s):  
Stéphanie Pommier ◽  
Marthe Gavioli ◽  
Eric Cascales ◽  
Roland Lloubès
Keyword(s):  
Escherichia Coli ◽  
Protein Complexes ◽  
Cell Envelope ◽  
Membrane Integrity ◽  
Binding Activity ◽  
Binding Motif ◽  
Cell Integrity ◽  
Outer Membranes

ABSTRACT The Tol-Pal proteins of the cell envelope of Escherichia coli are required for maintaining outer membrane integrity. This system forms protein complexes in which TolA plays a central role by providing a bridge between the inner and outer membranes via its interaction with the Pal lipoprotein. The Tol proteins are parasitized by filamentous bacteriophages and group A colicins. The N-terminal domain of the Ff phage g3p protein and the translocation domains of colicins interact directly with TolA during the processes of import through the cell envelope. Recently, a four-amino-acid sequence in Pal has been shown to be involved in Pal's interaction with TolA. A similar motif is also present in the sequence of two TolA partners, g3p and colicin A. Here, a mutational study was conducted to define the function of these motifs in the binding activity and import process of TolA. The various domains were produced and exported to the bacterial periplasm, and their cellular effects were analyzed. Cells producing the g3p domain were tolerant to colicins and filamentous phages and had destabilized outer membranes, while g3p deleted of three residues in the motif was affected in TolA binding and had no effect on cell integrity or colicin or phage import. A conserved Tyr residue in the colicin A translocation domain was involved in TolA binding and colicin A import. Furthermore, in vivo and in vitro coprecipitation analyses demonstrated that colicin A and g3p N-terminal domains compete for binding to TolA.

The Escherichia coli DNA translocase FtsK

2010 ◽  
Vol 38 (2) ◽  
pp. 395-398 ◽  
Author(s):  
David J. Sherratt ◽  
Lidia K. Arciszewska ◽  
Estelle Crozat ◽  
James E. Graham ◽  
Ian Grainge
Keyword(s):  
Escherichia Coli ◽  
Chromosome Segregation ◽  
Regulatory Domain ◽  
Biological Functions ◽  
Comparable Rate ◽  
Site Specific ◽  
Late Stages ◽  
Dna Translocase

Escherichia coli FtsK is a septum-located DNA translocase that co-ordinates the late stages of cytokinesis and chromosome segregation. Relatives of FtsK are present in most bacteria; in Bacillus subtilis, the FtsK orthologue, SpoIIIE, transfers the majority of a chromosome into the forespore during sporulation. DNA translocase activity is contained within a ~ 512-amino-acid C-terminal domain, which is divided into three subdomains: α, β and γ. α and β comprise the translocation motor, and γ is a regulatory domain that interacts with DNA and with the XerD recombinase. In vitro rates of translocation of ~ 5 kb·s−1 have been measured for both FtsK and SpoIIIE, whereas, in vivo, SpoIIIE has a comparable rate of translocation. Translocation by both of these proteins is not only rapid, but also directed by DNA sequence. This directionality requires interaction of the γ subdomain with specific 8 bp DNA asymmetric sequences that are oriented co-directionally with replication direction of the bacterial chromosome. The γ subdomain also interacts with the XerCD site-specific recombinase to activate chromosome unlinking by recombination at the chromosomal dif site. In the present paper, the properties in vivo and in vitro of FtsK and its relatives are discussed in relation to the biological functions of these remarkable enzymes.

scholarly journals The quantification of protein-protein interaction interfaces using solvent-excluded surface-defined properties

2018 ◽  
Author(s):  
Lincong Wang
Keyword(s):  
Protein Interaction ◽  
Protein Complexes ◽  
Protein Docking ◽  
Water Soluble ◽  
Surface Model ◽  
Protein Protein Interaction ◽  
Ring Model ◽  
Water Soluble Protein ◽  
Site Identification

AbstractProtein-protein interaction (PPI) is the cornerstone of nearly every biological process. During last forty years PPI interfaces have been investigated extensively both in vitro and in silico in order to understand both the strength and specificity of PPI. At least three different models, the buried surface model, the O-ring model and the rim- and-core model, have been proposed for PPI interface. However none of them provide much detail about PPI and a single model that reconciles them remains elusive. To identify common physical and geometrical features shared by various PPI interfaces we have analyzed several solvent-excluded surface (SES)-defined properties for a set of well-studied protein-protein complexes with crystal structures. Our analysis shows that the SES-defined properties for the interface atoms of a PPI partner are in general different from those for the surface atoms of a water-soluble protein. Most significantly we find that the partially-buried atoms of a PPI partner have unique SES-defined properties that set them well apart from either the buried atoms or the accessible atoms. Based on distinct SES-defined properties for the accessible, buried and partially-buried atoms shared by various PPI interfaces we propose a new model specified by a list of SES-defined properties shared by various PPI interfaces. Our model is quantitative in nature and should be useful for PPI site identification, protein-protein docking and structure-based design of chemicals targeting PPI.

scholarly journals Purification of complexes of nuclear oncogene p53 with rat and Escherichia coli heat shock proteins: in vitro dissociation of hsc70 and dnaK from murine p53 by ATP.

1988 ◽  
Vol 8 (3) ◽  
pp. 1206-1215 ◽  
Author(s):  
C F Clarke ◽  
K Cheng ◽  
A B Frey ◽  
R Stein ◽  
P W Hinds ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Protein Interactions ◽  
Protein Complexes ◽  
P53 Protein ◽  
Immunoaffinity Column ◽  
E Coli ◽  
Shock Proteins

Oligomeric protein complexes containing the nuclear oncogene p53 and the simian virus 40 large tumor antigen (D. I. H. Linzer and A. J. Levine, Cell 17:43-51, 1979), the adenovirus E1B 55-kilodalton (kDa) tumor antigen, and the heat shock protein hsc70 (P. Hinds, C. Finlay, A. Frey, and A. J. Levine, Mol. Cell. Biol. 7:2863-2869, 1987) have all been previously described. To begin isolating, purifying, and testing these complexes for functional activities, we have developed a rapid immunoaffinity column purification. p53-protein complexes are eluted from the immunoaffinity column by using a molar excess of a peptide comprising the epitope recognized by the p53 monoclonal antibody. This mild and specific elution condition allows p53-protein interactions to be maintained. The hsc70-p53 complex from rat cells is heterogeneous in size, with some forms of this complex associated with a 110-kDa protein. The maximum apparent molecular mass of such complexes is 660,000 daltons. Incubation with micromolar levels of ATP dissociates this complex in vitro into p53 and hsc70 110-kDa components. Nonhydrolyzable substrates of ATP fail to promote this dissociation of the complex. Murine p53 synthesized in Escherichia coli has been purified 660-fold on the same antibody affinity column and was found to be associated with an E. coli protein of 70 kDa. Immunoblot analysis with specific antisera demonstrated that this E. coli protein was the heat shock protein dnaK, which has extensive sequence homology with the rat hsc70 protein. Incubation of the immunopurified p53-dnaK complex with ATP resulted in the dissociation of the p53-dnaK complex as it did with the p53-hsc70 complex. This remarkable conservation of p53-heat shock protein interactions and the specificity of dissociation reactions suggest a functionally important role for heat shock proteins in their interactions with oncogene proteins.

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