Effect of acidic phospholipids on apolipoprotein binding by artificial lipid particles in vivo

1986 ◽  
Vol 64 (8) ◽  
pp. 836-846 ◽  
Author(s):  
Ming-For Tong ◽  
Arnis Kuksis
Keyword(s):  
Egg Yolk ◽  
Bovine Brain ◽  
Relative Strength ◽  
Minor Components ◽  
Qualitative And Quantitative ◽  
Egg Yolk Phosphatidylcholine ◽  
Unknown Protein ◽  
Lipid Particles ◽  
Acidic Phospholipids

Soybean triacylglycerol particles stabilized with soybean phosphatidylinositol (PI), bovine brain phosphatidylserine (PS), egg yolk phosphatidylcholine (PC) or mixtures of these acidic and neutral phospholipids were prepared with diameters ranging from 250 to 520 nm. Binding of apoproteins to the lipid particles was studied using the strategy of Connelly and Kuksis. The recoveries of the injected particles, which had undergone minimal changes in lipid composition, ranged from 57% for the PC-stabilized emulsions to 21% for the emulsions stabilized with PS and 8% for the emulsions stabilized with PI. The apoprotein (apo) composition of the recovered particles showed characteristic qualitative and quantitative differences. The particles stabilized with PI and PS or PI-phosphatidylethanolamine contained an unknown protein of molecular weight 117 000 (43–48%) and albumin (9–13%) as major components. The apoC-II, apoC-III, apoA-I, apoE, and apoA-IV were present as minor components in ratios that were the reverse of those seen for the PC-stabilized particles, which contained these proteins as major components. The relative strength of the binding of the proteins, which was determined by washing the particles with saline under standard conditions, also showed variations among the different particles and different apoproteins. The lipid particles stabilized with the acidic phospholipids had less total apoprotein and held it less tightly than the particles stabilized with PC. It is concluded that the binding of apoproteins by lipid particles stabilized with acidic phospholipids involves hydrophobic and ionic interactions, both of which may be physiologically important.

Effect of different neutral phospholipids on apolipoprotein binding by artificial lipid particles in vivo

1986 ◽  
Vol 64 (8) ◽  
pp. 826-835 ◽  
Author(s):  
Ming-for Tong ◽  
Arnis Kuksis
Keyword(s):  
Egg Yolk ◽  
Hepatic Uptake ◽  
Plasma Lipoproteins ◽  
Variable Component ◽  
Qualitative And Quantitative ◽  
Lipid Particles ◽  
Rapid Hydrolysis ◽  
A Minor ◽  
Hydrolysis Of

Soybean triacylglycerol particles, stabilized with egg yolk sphingomyelin (SPH), phosphatidylcholine (PC), phosphatidylethanolamine (PE), or PC–PE mixtures, with diameters ranging from 170 to 550 nm were prepared by sonication and isolated by ultracentrifugation. Binding of apoproteins to the lipid particles was studied in vivo using the strategy of Connelly and Kuksis. The recoveries of the injected particles, which had decreased in size and undergone minimal changes in lipid composition, ranged from 70% and 57% for SPH- and PC-stabilized particles to 14% for particles stabilized with egg yolk PC–PE mixtures. The apoprotein (apo) composition of the recovered particles showed qualitative and quantitative differences, which were affected by the number of washes during isolation. After four washes, the SPH and PC particles contained apoE, apoC-II, and apoC-III as major components and apoA-IV as minor components. In addition, all particles, except those stabilized with egg yolk PC, contained large amounts of albumin. In contrast to egg yolk PC, the dipalmitoyl PC particles bound albumin as a major component. The recovered PC-PE and PE particles were characterized by a relative decrease of apoC and greatly increased binding of albumin. The higher rate of clearance of the PE-containing particles was attributed to a relative absence of apoC-III, which is known to delay hepatic uptake of lipid particles containing it, and to a more rapid hydrolysis of PE by lipoprotein lipases. Since PE occurs as a minor and variable component of chylomicrons and plasma lipoproteins, the present observations are of physiological interest.

scholarly journals Sphingomyelin exhibits greatly enhanced protection compared with egg yolk phosphatidylcholine against detergent bile salts

2000 ◽  
Vol 41 (6) ◽  
pp. 916-924 ◽  
Author(s):  
Antonio Moschetta ◽  
Gerard P. vanBerge-Henegouwen ◽  
Piero Portincasa ◽  
Giuseppe Palasciano ◽  
Albert K. Groen ◽  
...  
Keyword(s):  
Bile Salts ◽  
Egg Yolk ◽  
Egg Yolk Phosphatidylcholine

Surface culture of Steinernema sp. on two solid media and their pathogenicity against Galleria mellonella

2021 ◽  
Vol 95 ◽  
Author(s):  
C.I. Cortés-Martínez ◽  
A.I. Rodríguez-Hernández ◽  
M.R. López-Cuellar ◽  
N. Chavarría-Hernández
Keyword(s):  
Galleria Mellonella ◽  
Egg Yolk ◽  
Infective Juvenile ◽  
Surface Culture ◽  
Solid Media ◽  
Significant Difference ◽  
Bacterial Lawn ◽  
The Mean

Abstract The use of native entomopathogenic nematodes as biocontrol agents is a strategy to decrease the environmental impact of insecticides and achieve sustainable agriculture crops. In this study, the effect of the surface culture of Steinernema sp. JAP1 over two solid media at 23–27°C on infective juvenile (IJ) production and pathogenicity against Galleria mellonella larvae were investigated. First, the bacterial lawn on the surface of the media with egg yolk (P2) or chicken liver (Cl) were incubated in darkness at 30°C for 48 and 72 h, and 100 surface-sterilized IJs were added. Four harvests were conducted within the next 35 days and the mean accumulated production was superior on Cl (210 × 103 IJs) than on P2 (135 × 103 IJs), but the productivity decreased up to 10% when the incubation time of the bacterial lawn was of 72 h. The mean pathogenicity of in vitro- and in vivo-produced IJs were of 47–64% and 31%, respectively. It is worth noting that none of the two solid media had a statistically significant difference in IJ pathogenicity. Considering that the maximum multiplication factor of IJs on solid media was 2108 and that the pathogenicity against G. mellonella was outstanding, Steinernema sp. has a good potential for in vitro mass production.

scholarly journals Screening of phospholipase A activity and its production by new actinomycete strains cultivated by solid-state fermentation

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3524 ◽  
Author(s):  
Priscila Sutto-Ortiz ◽  
María de los Angeles Camacho-Ruiz ◽  
Manuel R. Kirchmayr ◽  
Rosa María Camacho-Ruiz ◽  
Juan Carlos Mateos-Díaz ◽  
...  
Keyword(s):  
Solid State ◽  
Solid State Fermentation ◽  
High Throughput Screening ◽  
Egg Yolk ◽  
Specific Substrate ◽  
Phospholipase Activity ◽  
High Accumulation ◽  
Egg Yolk Phosphatidylcholine ◽  
Potential Applications ◽  
Routine Assay

Novel microbial phospholipases A (PLAs) can be found in actinomycetes which have been poorly explored as producers of this activity. To investigate microbial PLA production, efficient methods are necessary such as high-throughput screening (HTS) assays for direct search of PLAs in microbial cultures and cultivation conditions to promote this activity. About 200 strains isolated with selected media for actinomycetes and mostly belonging toStreptomyces(73%) andMicromonospora(10%) genus were first screened on agar-plates containing the fluorophore rhodamine 6G and egg yolk phosphatidylcholine (PC) to detect strains producing phospholipase activity. Then, a colorimetric HTS assay for general PLA activity detection (cHTS-PLA) using enriched PC (≈60%) as substrate and cresol red as indicator was developed and applied; this cHTS-PLA assay was validated with known PLAs. For the first time, actinomycete strains were cultivated by solid-state fermentation (SSF) using PC as inductor and sugar-cane bagasse as support to produce high PLA activity (from 207 to 2,591 mU/g of support). Phospholipase activity of the enzymatic extracts from SSF was determined using the implemented cHTS-PLA assay and the PC hydrolysis products obtained, were analyzed by TLC showing the presence of lyso-PC. Three actinomycete strains of theStreptomycesgenus that stood out for high accumulation of lyso-PC, were selected and analyzed with the specific substrate 1,2-α-eleostearoyl-sn-glycero-3-phosphocholine (EEPC) in order to confirm the presence of PLA activity in their enzymatic extracts. Overall, the results obtained pave the way toward the HTS of PLA activity in crude microbial enzymatic extracts at a larger scale. The cHTS-PLA assay developed here can be also proposed as a routine assay for PLA activity determination during enzyme purification,directed evolution or mutagenesis approaches. In addition, the production of PLA activity by actinomycetes using SSF allow find and produce novel PLAs with potential applications in biotechnology.

scholarly journals In vivo administration of anti-HIV hyper-immune eggs induces anti-anti-idiotypic antibodies against HIV gp120 peptides (fragments 308-331 and 421-438) and against HIV gp41 peptide (fragment 579-601) which have demonstrable protective capacity

2021 ◽  
Author(s):  
Angel Justiz-Vaillant ◽  
Belkis Ferrer-Cosme ◽  
Monica Fisher Smikle ◽  
Oliver Pérez
Keyword(s):  
Immune Responses ◽  
Hiv Infections ◽  
Egg Yolk ◽  
Laboratory Animals ◽  
Future Research ◽  
Protective Capacity ◽  
Antigen Binding Site ◽  
Hiv Antigen ◽  
Gp41 Peptide

AbstractIsolation of antibodies from the egg yolk of chickens is of particular interest as a source of specific antibodies for oral administration to prevent infections and use them as immunodiagnostic reagents. The use of birds in antibody production results in a reduction in the use of laboratory animals. Immunized chickens produce larger quantities of antibodies (2000 mg IgY/month) than rodents (200 mg IgG/month) in the laboratory. According to Jerne’s network theory, it is possible to produce an antibody against the antigen-binding site of another antibody. This study assessed the hypothesis that immunization with viral peptides (immunogens) could provide a potent immune response that could be evaluated in chicken eggs. Human immunodeficiency virus 1(HIV-1) is used as an immunogen. The second hypothesis was that an orally administered antibody stimulates the production of a complementary antibody, the so-called anti-idiotypic antibody, which can potentially be therapeutical. This study reports and analyzes the use of eggs as therapeutic agents. We wanted to test the hypothesis that feeding chicks with hyperimmune eggs stimulates the production of anti-anti-idiotypic antibodies that neutralize the original HIV antigen fragments 308-331 or 421-438 of gp120 or fragment 579-601 of gp41. Future research could entail an anti-idiotype strategy for prophylactic vaccines. It is vital to note that it may need an anti-idiotype response to prime immunity against an HIV viral epitope, which may be used as a secondary element. The use of anti-idiotype immune responses in infected individuals may shift the balance of the immune system, allowing the organism to manage HIV infection. Therefore, it may be an avenue for immunotherapy to improve the fight against HIV infections. However, more studies and clinical trials are required to demonstrate similar human immune responses as observed in birds.

The complex structure of polyhydroxybutyrate (PHB) granules: four orthologous and paralogous phasins occur in Ralstonia eutropha

Microbiology ◽  
2004 ◽  
Vol 150 (7) ◽  
pp. 2301-2311 ◽  
Author(s):  
Markus Pötter ◽  
Helena Müller ◽  
Frank Reinecke ◽  
Roman Wieczorek ◽  
Florian Fricke ◽  
...  
Keyword(s):  
Ralstonia Eutropha ◽  
Azotobacter Vinelandii ◽  
Production Systems ◽  
Complex Structure ◽  
Pcr Analysis ◽  
Unknown Protein ◽  
Considerable Impact ◽  
Unexpected Findings

Analysis of the genome sequence of the polyhydroxyalkanoate- (PHA) accumulating bacterium Ralstonia eutropha strain H16 revealed three homologues (PhaP2, PhaP3 and PhaP4) of the phasin protein PhaP1. PhaP1 is known to constitute the major component of the layer at the surface of poly(3-hydroxybutyrate), poly(3HB), granules. PhaP2, PhaP3 and PhaP4 exhibited 42, 49 and 45 % identity or 61, 62 and 63 % similarity to PhaP1, respectively. The calculated molecular masses of PhaP1, PhaP2, PhaP3 and PhaP4 were 20·0, 20·2, 19·6 and 20·2 kDa, respectively. RT-PCR analysis showed that phaP2, phaP3 and phaP4 were transcribed under conditions permissive for accumulation of poly(3HB). 2D PAGE of the poly(3HB) granule proteome and analysis of the detected proteins by MALDI-TOF clearly demonstrated that PhaP1, PhaP3 and PhaP4 are bound to the poly(3HB) granules in the cells. PhaP3 was expressed at a significantly higher level in PhaP1-negative mutants. Occurrence of an unknown protein with an N-terminal amino-acid sequence identical to that of PhaP2 in crude cellular extracts of R. eutropha had previously been shown by others. Although PhaP2 could not be localized in vivo on poly(3HB) granules, in vitro experiments clearly demonstrated binding of PhaP2 to these granules. Further analysis of complete or partial genomes of other poly(3HB)-accumulating bacteria revealed the existence of multiple phasin homologues in Ralstonia solanacearum, Burkholderia fungorum and Azotobacter vinelandii. These new and unexpected findings should affect our current models of PHA-granule structure and may also have a considerable impact on the establishment of heterologous production systems for PHAs.

scholarly journals Phytochemical Analysis of Podospermum and Scorzonera n-Hexane Extracts and the HPLC Quantitation of Triterpenes

Molecules ◽  
2018 ◽  
Vol 23 (7) ◽  
pp. 1813 ◽  
Author(s):  
Özlem Bahadır-Acıkara ◽  
Serkan Özbilgin ◽  
Gülcin Saltan-İşcan ◽  
Stefano Dall’Acqua ◽  
Veronika Rjašková ◽  
...  
Keyword(s):  
Hplc Method ◽  
Phytochemical Analysis ◽  
Qualitative And Quantitative Analysis ◽  
Qualitative And Quantitative ◽  
Aerial Parts ◽  
Hexane Extracts ◽  
Lupeol Acetate ◽  
Anti Inflammatory

Previously tested n-hexane extracts of the Scorzonera latifolia showed promising bioactivity in vivo. Because triterpenes could account for this activity, n-hexane extracts were analyzed by HPLC to identify and quantify the triterpenes as the most abundant constituents. Other Scorzonera and Podospermum species, potentially containing triterpenic aglycones, were included in the study. An HPLC method for simultaneous determination of triterpene aglycones was therefore developed for analysis of Podospermum and Scorzonera species. n-Hexane extracts of root and aerial parts of S. latifolia, ten other Scorzonera species and two Podospermum species were studied to compare the content of triterpenes. HPLC was used for the qualitative and quantitative analysis of α-amyrin, lupeol, lupeol acetate, taraxasteryl acetate, 3-β-hydroxy-fern-7-en-6-one acetate, urs-12-en-11-one-3-acetyl, 3-β-hydroxy-fern-8-en-7-one acetate, and olean-12-en-11-one-3-acetyl. Limits of detection and quantification were determined for each compound. HPLC fingerprinting of n-hexane extracts of Podospermum and Scorzonera species revealed relatively large amounts of triterpenes in a majority of investigated taxa. Lupeol, lupeol acetate, and taraxasteryl acetate were found in a majority of the species, except S. acuminata. The presence of α-amyrin, 3β-hydroxy-fern-7-en-6-one-acetate, urs-12-en-11-one-3-acetyl, 3β-hydroxy-fern-8-en-7-one-acetate, and olean-12-en-11-one-3-acetyl was detected in varying amounts. The triterpene content could correlate with the analgesic and anti-inflammatory activity of Scorzonera, which was previously observed and Scorzonera species that have been determined to contain triterpenes in large amounts and have not yet been tested for their analgesic activity should be tested for their potential analgesic and anti-inflammatory potential. The presented HPLC method can be used for analysis of triterpene aglycones, for example dedicated to chemosystematic studies of the Scorzonerinae.

Selection of splice sites in pre-mRNAs with short internal exons

1991 ◽  
Vol 11 (12) ◽  
pp. 6075-6083
Author(s):  
Z Dominski ◽  
R Kole
Keyword(s):  
Alternative Pathway ◽  
Globin Gene ◽  
Relative Strength ◽  
Splice Sites ◽  
Polypyrimidine Tract ◽  
Splice Site Selection ◽  
Nuclear Extracts ◽  
Internal Exon

Model pre-mRNAs containing two introns and three exons, derived from the human beta-globin gene, were used to study the effects of internal exon length on splice site selection. Splicing was assayed in vitro in HeLa nuclear extracts and in vivo during transient expression in transfected HeLa cells. For substrates with internal exons 87, 104, and 171 nucleotides in length, in vitro splicing proceeded via a regular splicing pathway, in which all three exons were included in the spliced product. Primary transcripts with internal exons containing 23, 29, and 33 nucleotides were spliced by an alternative pathway, in which the first exon was joined directly to the third one. The internal exon was missing from the spliced product and together with two flanking introns was included in a large lariat structure. The same patterns of splicing were retained when transcripts containing 171-, 33-, and 29-nucleotide-long internal exons were spliced in vivo. A transcript containing a 51-nucleotide-long exon was spliced in vitro via both pathways but in vivo generated only a correctly spliced product. Skipping of short internal exons was reversed both in vitro and in vivo when purines in the upstream polypyrimidine tract were replaced by pyrimidines. The changes in the polypyrimidine tract achieved by these substitutions led in vitro to complete (transcripts containing 28 pyrimidines in a row) or partial (transcripts containing 15 pyrimidines in a row) restoration of a regular splicing pathway. Splicing in vivo of these transcripts led exclusively to the spliced product containing all three exons. These results suggest that a balance between the length of the uninterrupted polypyrimidine tract and the length of the exon is an important determinant of the relative strength of the splice sites, ensuring correct splicing patterns of multiintron pre-mRNAs.

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