scholarly journals Uptake and degradation of iodine-labelled chylomicron remnant particles by monolayers of rat hepatocytes

1978 ◽  
Vol 174 (3) ◽  
pp. 827-838 ◽  
Author(s):  
C H Florén ◽  
A Nilsson
Keyword(s):  
Protein A ◽  
Sodium Dodecyl ◽  
Rat Hepatocytes ◽  
Relative Increase ◽  
Cell Protein ◽  
Chylomicron Remnant ◽  
Protein Mass ◽  
Apolipoprotein A ◽  
Chylomicron Remnants ◽  
Protein Moiety

1. Rat chylomicrons were labelled with 125I with 69–72% of the iodine in the protein moiety. Less than 1 nmol of iodine was incorporated per nmol of protein. Of the peptide radioactivity 44–56% was in apolipoprotein A-1, 30–40% in the C peptides and 11–15% in apolipoprotine B. The arginine-rich peptide, which accounted for about 14% of the chylomicron protein mass as determined by scanning of sodium dodecyl sulphate-polyacrylamide gels, contained very little radioactivity. 2. Chylomicron remnants generated with postheparin plasma from iodine-labelled chylomicrons showed a relative increase in the percentage of the arginine-rich peptide (76–90% of the apolipoprotein mass according to gel scanning). The major portion of the peptide iodine label was present in apolipoprotein A-1 (43–57%), B (22–32%) and C peptides (17–35%). 3. When iodine-labelled chylomicron remnants were added to rat hepatocytes in primary culture, labelled peptides were taken up and degraded by the hepatocytes by a saturable process. The Vmax. for the uptake was calculated to the 300ng of protein/h per mg of cell protein and the apparent Km as 7.7 microgram of protein/mg of cell protein. A larger proportion of the 125I-labelled lipids of the remnants (mainly polar lipids) was taken up. This suggest that these may also enter the cells by a mechanism that does not involve particulate uptake, such as phospholipid exchange. 4. The degradation of labelled peptides was inhibited by colchicine, concanavalin A, chloroquine and NH4Cl, which also inhibit degradation of the cholesteryl ester portion. All these drugs exerted their inhibition mainly after the uptake of labelled peptide. No degradation occurred at 4 degrees C, and also the uptake was markedly decreased. 5. The uptake of labelled chylomicron remnant peptide was 77 times as effective as that of labelled sucrose, which is likely to be taken up randomly by pinocytosis.

scholarly journals Exercise-mitigated gender-based differences in aging: From genetic alterations to heart performance

2020 ◽  
Author(s):  
Denise Börzsei ◽  
Daniel Priksz ◽  
Renáta Szabó ◽  
Mariann Bombicz ◽  
Zoltán Karácsonyi ◽  
...  
Keyword(s):  
Heart Function ◽  
Protein A ◽  
Genetic Alterations ◽  
Functional Adaptation ◽  
Voluntary Exercise ◽  
Female Rats ◽  
Cell Protein ◽  
Calcium Balance ◽  
Male And Female Rats ◽  
Heart Performance

The prevalence of cardiovascular diseases dramatically increases with age, therefore striving to maintain a physiological heart function became particularly important. We aimed to study the voluntary exercise evoked cardioprotective effects in aged male and female rats, from genetic alterations to changes in heart performance. We divided 20-month-old female and male Wistar rats to control and running groups. After the 12-week-long experimental period, echocardiographic measurements were performed. Afterwards, hearts were either removed for biochemical measurements or mounted into a Langendorff-perfusion system to detect infarct size. The following genes and their proteins were analyzed from heart: catechol-O-methyltransferase (Comt), endothelin-1 (Esm1), Purkinje cell protein-4 (Pcp4), and osteoglycin (Ogn). Recreational exercise caused functional improvements; however, changes were more prominent in males. Cardiac expression of Comt and Ogn were reduced as a result of exercise in aged males, while Pcp4 and Esm1 showed a marked overexpression, along with a markedly improved diastolic function. The key result of this study is that exercise enhanced the expression of the Pcp4 gene and protein, a recently described regulator of calcium balance in cardiomyocytes, and suppressed Comt and Ogn gene expression, that has been associated with impaired cardiac function. In addition, as a result of exercise, a significant improvement was observed in the size of infarct elicited by left anterior descending coronary artery occlusion. Our results clearly show that age and sex-dependent changes were both apparent in key proteins linked to cardiovascular physiology. Exercise-moderated fundamental genetic alterations may have contributed to the functional adaptation of the heart.

scholarly journals Analysis with specific polyclonal antiserum indicates that the E1A-associated 300-kDa product is a stable nuclear phosphoprotein that undergoes cell cycle phase-specific modification.

1991 ◽  
Vol 11 (11) ◽  
pp. 5389-5397 ◽  
Author(s):  
P Yaciuk ◽  
E Moran
Keyword(s):  
Cell Cycle ◽  
Cell Function ◽  
Rat Kidney ◽  
Sodium Dodecyl ◽  
Polyclonal Antiserum ◽  
Cell Protein ◽  
Cycle Phase ◽  
Resting Cells ◽  
Kidney Cells ◽  
Nuclear Phosphoprotein

Binding of a 300-kDa host cell protein (p300) is tightly correlated with the ability of the adenovirus E1A products to induce quiescent baby rat kidney cells to proliferate. We have generated rabbit polyclonal antibodies against p300 to characterize this protein further. We have found p300 to be a nuclear phosphoprotein that is actively synthesized in both quiescent and proliferating baby rat kidney cells. In partially purified mitotic cell populations, we observe a form of p300 with decreased electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels that shares a nearly identical partial proteolytic digest pattern with p300. The slower-migrating form of p300 is greatly reduced by treating immune complexes with potato acid phosphatase. The relative stability and presence of p300 even in resting cells suggests that p300 has a basal cell function, but the appearance of differentially modified forms during the cell cycle suggests the possibility that p300 function is modulated specifically in growing cells.

Solubilization and immunoprecipitation of 125I-labelled antigens from Plasmodium knowlesi schizont-infected erythrocytes using non-ionic, anionic and zwitterionic detergents

Parasitology ◽  
1984 ◽  
Vol 88 (1) ◽  
pp. 27-36 ◽  
Author(s):  
R. J. Howard ◽  
J. W. Barnwell
Keyword(s):  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Plasmodium Knowlesi ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Ionic Detergents ◽  
Class A ◽  
Anionic Detergents ◽  
Triton X

SUMMARYPlasmodium knowlesi malaria-infected erythrocytes were radio-iodinated and several non-ionic, anionic and zwitterionic detergents were compared in their capacity to extract the labelled membrane proteins. The use of these detergents for antigen identification was tested by immunoprecipitation, after addition of Triton X-100 to some detergent extracts, using hyperimmune monkey antiserum and protein A-Sepharose. 125I-labelled antigens were specifically immunoprecipitated with all detergents tested, including the anionic detergents sodium dodecyl sulphate (SDS), deoxycholate and cholate; the zwitterions Zwittergent-312 and -314, CHAPS and Empigen BB, as well as several non-ionic detergents. The SDS-polyacrylamide gel electrophoresis patterns of 125I-labelled antigens varied after extraction with different detergents, there being no consistent pattern for detergents of a particular class. A total of 14 125I-labelled antigens were identified, 11 of them using Triton X-100. Some minor antigens identified with Triton X-100 were immunoprecipitated in greater amount after extraction in other detergents. Most importantly, two antigens Mr 200000 and 180000 were detected only after extraction with deoxycholate or SDS.

Identification of pilin pools in the membranes of Pseudomonas aeruginosa

1982 ◽  
Vol 152 (2) ◽  
pp. 687-691
Author(s):  
T H Watts ◽  
E A Worobec ◽  
W Paranchych
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Gel Electrophoresis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Wild Type ◽  
Outer Membranes

The proteins of purified inner and outer membranes obtained from Pseudomonas aeruginosa strains PAK and PAK/2Pfs were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and treated with antiserum raised against pure pili. Bound antipilus antibodies were visualized by reaction with 125I-labeled protein A from Staphylococcus aureus. The results showed that there are pools of pilin in both the inner and outer membranes of P. aeruginosa and that the pool size in the multipiliated strain is comparable with that of the wild-type strain.

Abscisic acid and low temperature induced polypeptide changes in alfalfa (Medicago sativa) cell suspension cultures

1986 ◽  
Vol 64 (11) ◽  
pp. 2758-2763 ◽  
Author(s):  
Albert J. Robertson ◽  
Lawrence V. Gusta
Keyword(s):  
Abscisic Acid ◽  
Low Temperature ◽  
Medicago Sativa ◽  
Cell Suspension ◽  
Sodium Dodecyl ◽  
Cell Suspension Cultures ◽  
Suspension Cultures ◽  
Total Cell ◽  
Cell Protein ◽  
Major Protein

Changes in extracellular, cellular, and subcellular proteins during abscisic acid and low temperature induced cold hardening of alfalfa (Medicago sativa L. cv. Wisconsin 22C) cell suspension cultures were investigated by sodium dodecyl sulphate polyacrylamide gel electrophoresis. Extracellular proteins from 4- to 6-day-old ABA and low temperature grown alfalfa cells showed decreased electrophoretic mobilities, lacked a 190-kDa glycoprotein, and had reduced amounts of four other polypeptides. In total cell protein analyses, a 42-kDa protein was enriched in both ABA and low temperature treated alfalfa cells. Several proteins increased or induced by exogenous ABA treatment were identified in the extracellular (12.5 and 13 to 15 kDa), total cell and cell wall (24 kDa), and soluble (20, 37, and 41 kDa) fractions. However, no major protein changes were resolved by one-dimensional electrophoretic analyses of crude membrane proteins.

Preliminary Treatment of Urinary Proteins Improves Electrophoretic Analysis and Immunodetection

1992 ◽  
Vol 38 (6) ◽  
pp. 860-863 ◽  
Author(s):  
J M Verdier ◽  
B Dussol ◽  
P Dupuy ◽  
Y Berland ◽  
J C Dagorn
Keyword(s):  
Protein Pattern ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Complex Mixture ◽  
Electrophoretic Analysis ◽  
Renal Physiology ◽  
Preliminary Treatment ◽  
Urinary Proteins ◽  
Simple Method ◽  
Electrophoretic Migration

Abstract Analysis of urinary protein composition is an important tool in studies on renal physiology and physiopathology. Urine is, however, a complex mixture containing, besides protein, a variety of compounds such as salts, peptides, oligosaccharides, and glycosaminoglycans. Some of these compounds interfere with the electrophoretic migration of protein in sodium dodecyl sulfate-polyacrylamide gels and prevent correct analysis of the protein pattern. We describe a simple method for extracting urinary proteins that considerably improves their electrophoretic migration and subsequent immunodetection. This treatment involves ammonium sulfate fractionations (for precipitating proteins), EDTA (for inhibiting protein aggregation), and HCl hydrolysis (for removing glycosylaminoglycans). Recovery during extraction was found to be almost quantitative for total protein and three representative proteins: albumin, alpha 1-glycoprotein acid, and beta 2-microglobulin.

scholarly journals Red cell proteins. I. Two-dimensional mapping of human erythrocyte lysate proteins

Blood ◽  
1979 ◽  
Vol 53 (6) ◽  
pp. 1121-1132 ◽  
Author(s):  
JJ Edwards ◽  
NG Anderson ◽  
SL Nance ◽  
NL Anderson
Keyword(s):  
Human Erythrocyte ◽  
Sodium Dodecyl ◽  
Deae Cellulose ◽  
Cell Protein ◽  
Red Cell ◽  
Two Dimensional ◽  
Two Dimensional Electrophoresis ◽  
Mapping Techniques ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Dimensional Electrophoresis

Abstract Human erythrocyte lysate proteins were resolved into over 250 discrete spots by two-dimensional electrophoresis using isoelectric focusing in the first dimension and electrophoresis in the presence of sodium dodecyl sulfate, (SDS) in the second. The overwhelming excess of hemoglobin has made such analyses difficult in the past. However, with the ISO-DALT two-dimensional electrophoresis system, large numbers of red cell proteins can be mapped in the presence of hemoglobin. When hemoglobin and several other major proteins are removed by adsorption to DEAE-cellulose, additional minor components are seen, giving a total of over 275. With the use of purified preparations, the map positions of five cell enzymes or their subunits were determined: pyruvate kinase, catalase, glucose-6-phosphate dehydrogenase, hypoxanthine phosphoribosyltransferase, and carbonic anhydrase. The mapping techniques described complement and extend those traditionally used to find human red cell protein variants.

scholarly journals Protein Engineering of Wzc To Generate New Emulsan Analogs

2007 ◽  
Vol 73 (12) ◽  
pp. 4020-4028 ◽  
Author(s):  
Hanna Dams-Kozlowska ◽  
David L. Kaplan
Keyword(s):  
Molecular Weight ◽  
Chain Length ◽  
Genetic Manipulation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Point Mutations ◽  
Biological Properties ◽  
Feeding Strategies ◽  
Exogenous Fatty Acid

ABSTRACT Acinetobacter venetianus Rag1 produces an extracellular, polymeric lipoheteropolysaccharide termed apoemulsan. This polymer is putatively produced via a Wzy-dependent pathway. According to this model, the length of the polymer is regulated by polysaccharide-copolymerase (PCP) protein. A highly conserved proline and glycine motif was identified in all members of the PCP family of proteins and is involved in regulation of polymer chain length. In order to control the structure of apoemulsan, defined point mutations in the proline-glycine-rich region of the apoemulsan PCP protein (Wzc) were introduced. Modified wzc variants were introduced into the Rag1 genome via homologous recombination. Stable chromosomal mutants were confirmed by Southern blot analysis. The molecular weight of the polymer was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Five of the eight point mutants produced polymers having molecular weights higher than the molecular weight of the polymer produced by the wild type. Moreover, four of these five polymers had modified biological properties. Replacement of arginine by leucine (R418L) resulted in the most significant change in the molecular weight of the polymer. The R418L mutant was the most hydrophilic mutant, exhibiting decreased adherence to polystyrene, and inhibited biofilm formation. The results described in this report show the functional effect of Wzc modification on the molecular weight of a high-molecular-weight polysaccharide. Moreover, in the present study we developed a genetic system to control polymerization of apoemulsan. The use of selective exogenous fatty acid feeding strategies, as well as genetic manipulation of sugar backbone chain length, is a promising new approach for bioengineering emulsan analogs.

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