Changes In Membrane Physicochemical Properties Of Hyperreactive Platelets In Estrogen-Treated Prostatic Cancer

1981 ◽  
Author(s):  
S M Jung ◽  
I Isohisa ◽  
K Kinoshita ◽  
H Yamazaki
Keyword(s):  
Sialic Acid ◽  
Physicochemical Properties ◽  
Electrophoretic Mobility ◽  
Prostatic Cancer ◽  
Polyacrylamide Gel Electrophoresis ◽  
Prostatic Hypertrophy ◽  
Factors Affecting ◽  
Glycoprotein I ◽  
Electrophoretic Mobilities

Aggregation and physicochemical properties of platelets from 5 prostatic cancer patients, 20 prostatic cancer on estrogen (300 mg stilbesterol diphosphate/day p.o.), 16 of the same patients given 300 mg aspirin/day, 19 prostatic hypertrophy and 21 aged males were examined. Prostatic cancer on estrogen show higher platelet aggregabilities towards ADP, adrenaline, and collagen than the other groups tested. Mean platelet electrophoretic mobilities were determined by the automatic Laser Zee 3000 system and the value for prostatic cancer on estrogen was -1.632±0.0182 um/sec/V/cm (mean ±SE). significantly higher (p <0.001)than the values for prostatic cancer not on estrogen, prostatic hypertrophy, and aged males with values of -1.534±0.0328, -1.526±0.009, and -1.535±0.012 um/sec/V/cm respectively. Electrophoretic mobility was a linear function of whole platelet sialic acid (r=0.96,n=66). The result suggests that the electrophoretic mobility of platelets was defined by sialic acid amount and there may be a selective consumption of platelets with less surface negative charge in prostatic cancer on estrogen. Membrane glycoprotein patterns determined by SDS-polyacrylamide gel electrophoresis showed prostatic cancer on estrogen to have lower glycoprotein I and higher glycoprotein IV than other groups (p < 0.05). There may be factors affecting changes on platelet membranes or population in such patients. Prostatic cancer on estrogen and aspirin had increased mobilities in 14 out of 16 cases with loss of linear relationship between mobility but no changes in sialic acid or glycoprotein pattern from those before aspirin. In vitro incubation of PRP with aspirin resulted in dose dependent increase in mobility with concomitant aggregability loss. The effect suggests another kind of inhibitory mechanism on platelet aggregation in addition to cyclooxygenase inhibition.

Partial resialylation of human asialotransferrin types 1 and 2 in the rat

1984 ◽  
Vol 62 (9) ◽  
pp. 853-858 ◽  
Author(s):  
Erwin Regoeczi ◽  
Paul A. Chindemi ◽  
Maria T. Debanne
Keyword(s):  
Sialic Acid ◽  
Electrophoretic Mobility ◽  
Acid Content ◽  
Sialic Acid Content ◽  
Small Doses ◽  
Type 3 ◽  
Time Type

125I-labeled asialotransferrin types 1 and 2 were administered in small doses to rats. The protein still in the plasma after 1–12 h was partially repurified and electrophoresed at pH 8.1, together with a transferrin standard that is composed of all six forms of the protein with respect to sialic acid content. The electrophoretic mobility of both asialotransferrins increased with time, type 2 being affected sooner than type 1. The changed mobility was due to increased electronegativity that was fully reversible by treatment of the samples with neuraminidase, thus identifying the underlying cause as partial resialylation. Asialotransferrin incubated in vitro with serum, plasma, or whole blood for 16 h exhibited no change in electrophoretic mobility. In conjunction with an earlier study on asialotransferrin type 3, it was found that the apparent speeds of resialylation of the three asialotransferrins were in the same order as their affinities for the asialoglycoprotein-binding hepatic lectin. This suggests the involvement of an endo- rather than of an ecto-transferase. Transfer of 59Fe from asialotransferrins to the liver was used to monitor the frequency of hepatocyte–asialotransferrin interactions. Iron deposition in the liver took place much more rapidly than the appearance of detectable quantities of partially resialylated asialotransferrin molecules in the circulation. It is concluded that each asialotransferrin molecule probably undergoes several passages through the hepatocyte before its glycans become modified.

Molecular basis for heterogeneity of the human p53 protein

1986 ◽  
Vol 6 (12) ◽  
pp. 4650-4656
Author(s):  
N Harris ◽  
E Brill ◽  
O Shohat ◽  
M Prokocimer ◽  
D Wolf ◽  
...  
Keyword(s):  
Electrophoretic Mobility ◽  
P53 Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Antigenic Determinants ◽  
Sequence Difference ◽  
Cdna Clones ◽  
Protein Species ◽  
Free System ◽  
The Individual

The human p53 tumor antigen comprises several physically distinct proteins. Two p53 proteins, separable by polyacrylamide gel electrophoresis, are expressed by the human transformed cell line SV-80. The individual cDNAs which code for these proteins were isolated and constructed into the SP6 transcription vector. The proteins encoded by these clones were identified by in vitro transcription with the SP6 vector and translation in a cell-free system. p53-H-1 and p53-H-19 cDNA clones code for the faster- and slower-migrating p53 protein species, respectively, of SV-80. The in vitro-expressed proteins of p53-H-1 and p53-H-19 had the same antigenic determinants and were structurally indistinguishable from their in vivo counterparts. By expressing defined restricted cDNA fragments in vitro, the region of heterogeneity between the respective cDNAs was located at the 5' end of the cDNAs. Exchanging the 5' fragments of interest and expressing the chimeric clones in vitro confirmed that the DNA heterogeneity was responsible for the difference in the electrophoretic mobility of these proteins. The sequences of the two cDNAs revealed a single base pair difference (G versus C) in the coding region of the clones. This sequence difference resulted in an arginine being coded for in clone p53-H-1 and a proline being coded for at the equivalent position in clone p53-H-19. This variation accounted for the change in the electrophoretic mobility of the individual p53 protein species.

Serum Catalase: Reversibly Formed Charge Isoform of Erythrocyte Catalase

1991 ◽  
Vol 37 (12) ◽  
pp. 2043-2047 ◽  
Author(s):  
László Góth
Keyword(s):  
Electrophoretic Mobility ◽  
Serum Proteins ◽  
Binding Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Permeation Chromatography ◽  
Electrophoretic Mobilities ◽  
Erythrocyte Catalase ◽  
Gel Permeation ◽  
Human Sera ◽  
Molecular Masses

Abstract The different electrophoretic mobilities of erythrocyte and serum catalase (EC 1.11.1.6) were confirmed and the causes responsible for their differences were examined. The presence of a catalase-binding protein in serum that could form a complex with erythrocyte catalase was excluded by incubating serum proteins with erythrocyte catalase. No new unequivocal catalase bands representing a catalase-binding protein were detected. The erythrocyte and serum catalase proved to be charge isoforms: their molecular masses, estimated by gel permeation chromatography or polyacrylamide gel electrophoresis in a nondenaturing system, were very similar, whereas their electrophoretic mobilities were different. Assay of serum catalase by gel permeation and hydrophobic chromatography yielded a product with the same electrophoretic mobility as that of erythrocyte catalase. Different dilution of erythrocyte catalase with human sera led to a gradual decrease of its mobility, 20-fold or greater dilution yielding the same results as for serum catalase. Similarly, when serum catalase was diluted 20-fold or more with 60 mmol/L phosphate buffer, it migrated similarly to erythrocyte catalase. I detected no effect of dialyzable serum ligands, NADPH, or protection of SH groups on the electrophoretic mobility of either catalase isoform. I conclude that formation of charge isoforms of catalase is caused by a reversible, conformational modification due to matrix effect of serum.

scholarly journals Structural glycoproteins from rabbit aortic media

1983 ◽  
Vol 211 (1) ◽  
pp. 257-265 ◽  
Author(s):  
M Moczar ◽  
B Phan-Dinh-Tuy ◽  
E Moczar ◽  
L Robert
Keyword(s):  
Amino Acids ◽  
Polyacrylamide Gel Electrophoresis ◽  
Guanidinium Chloride ◽  
Molecular Weights ◽  
Glycoprotein I ◽  
Aortic Media ◽  
Aortic Intima ◽  
Chloroform Methanol

Rabbit aortic intima-media fragments were incubated with [14C]mannose and [3H]fucose for 6 h to detect glycoproteins synthesized in situ. The radioactively labelled and the non-labelled samples were extracted with 0.2 mM-CaCl2/0.5 mM-dithiothreitol/0.5 mM-ATP and chloroform/methanol/water (4:4:1, by vol.). The delipidated residue was extracted with 5 M-guanidinium chloride/0.05 M-dithiothreitol/0.1 M-Tris/0.4% Na2EDTA, pH 7.5, before (extract 1) and after hydrolysis with collagenase (extract 2). The proteins in extracts 1 and 2 were S-carboxamidomethylated and separated by molecular-sieve chromatography, polyacrylamide-gel electrophoresis and isoelectric focusing in sucrose gradients in urea. The apparent molecular weights of glycoproteins were 36 000 (glycoprotein I) from extract 1, 50 000 (glycoprotein II) and 130 000 (glycoprotein III) from extract 2. The molecular weights of the non-labelled and radioactively labelled glycoproteins were identical. Glycoproteins I, II and III contain large amounts of polar amino acids and methionine. They contain neither hydroxyproline nor 3-methylhistidine. A hydroxyproline-containing component of 160 000-apparent-mol.wt. relatively rich in polar amino acids and labelled with incorporated sugars was isolated from extract 1. The incorporation in vitro of radioactive sugars into glycoproteins I, II, III and collagenous glycoproteins indicates that they are synthesized in the surviving aorta by the smooth-muscle cells.

scholarly journals Identification of a Novel Sialic Acid Transporter in Haemophilus ducreyi

2005 ◽  
Vol 73 (10) ◽  
pp. 6727-6735 ◽  
Author(s):  
Deborah M. B. Post ◽  
Rachna Mungur ◽  
Bradford W. Gibson ◽  
Robert S. Munson
Keyword(s):  
Sialic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Sialic Acid Residue ◽  
Haemophilus Ducreyi ◽  
Acid Uptake ◽  
Terminal Galactose ◽  
Genes Encoding ◽  
Acid Transporter

ABSTRACT Haemophilus ducreyi, the causative agent of chancroid, produces a lipooligosaccharide (LOS) which terminates in N-acetyllactosamine. This glycoform can be further extended by the addition of a single sialic acid residue to the terminal galactose moiety. H. ducreyi does not synthesize sialic acid, which must be acquired from the host during infection or from the culture medium when the bacteria are grown in vitro. However, H. ducreyi does not have genes that are highly homologous to the genes encoding known bacterial sialic acid transporters. In this study, we identified the sialic acid transporter by screening strains in a library of random transposon mutants for those mutants that were unable to add sialic acid to N-acetyllactosamine-containing LOS. Mutants that reacted with the monoclonal antibody 3F11, which recognizes the terminal lactosamine structure, and lacked reactivity with the lectin Maackia amurensis agglutinin, which recognizes α2,3-linked sialic acid, were further characterized to demonstrate that they produced a N-acetyllactosamine-containing LOS by silver-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis and mass spectrometric analyses. The genes interrupted in these mutants were mapped to a four-gene cluster with similarity to genes encoding bacterial ABC transporters. Uptake assays using radiolabeled sialic acid confirmed that the mutants were unable to transport sialic acid. This study is the first report of bacteria using an ABC transporter for sialic acid uptake.

ROLE OF GALACTOSE IN THE ANTIGENIC PROPERTIES OF THYROGLOBULIN

1976 ◽  
Vol 83 (1) ◽  
pp. 93-98 ◽  
Author(s):  
F. Monaco ◽  
M. Andreoli
Keyword(s):  
Sialic Acid ◽  
Thyroid Nodule ◽  
Electrophoretic Mobility ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Human Thyroid ◽  
Antigenic Properties ◽  
Beta Galactosidase

ABSTRACT Thyroglobulins (TG) from a "hot" human thyroid nodule and from Fisher rats have been purified and the effects of progressive removal of sialic acid and galactose on the immunoreactive properties of the proteins were studied. Terminal sialic acid and galactose were released by stepwise hydrolysis with neuraminidase and beta-galactosidase. Agalacto-TG shows a slower electrophoretic mobility than native TG, but in polyacrylamide gel electrophoresis and immunoelectrophoresis it migrates in the same position as asialo-TG. In immunodiffusion agalacto-TG forms a spur with native TG and asialo-TG when tested against anti 19S native TG or anti-asialo-TG sera. It is thus shown that galactose in the terminal environment of the oligosaccharide chains of thyroglobulin is essential for the structural groups involved in the antigenic properties of thyroglobulin.

scholarly journals Comparative effects of dry-aging and wet-aging on physicochemical properties and digestibility of Hanwoo beef

2020 ◽  
Vol 33 (3) ◽  
pp. 501-505 ◽  
Author(s):  
Ji-Han Kim ◽  
Tae-Kyung Kim ◽  
Dong-Min Shin ◽  
Hyun-Wook Kim ◽  
Young-Boong Kim ◽  
...  
Keyword(s):  
Physicochemical Properties ◽  
Shear Force ◽  
Myosin Light Chain ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
In Vitro Digestibility ◽  
Fragmentation Index ◽  
Effects Of Aging ◽  
Water Holding

Objective: The purpose of this study was to investigate the effects of aging methods (AM) i.e. dry-aging (DA) and wet-aging (WA) on the physicochemical properties and in vitro digestibility of proteins in beef short loin.Methods: Short loins (M. longissmus lumborum), were trimmed and boned-out on the fifth day postmortem, from a total of 18 Hanwoo, which were purchased from a commercial slaughterhouse. Short loins were separated randomly grouped into one of the three treatments: control, WA (1°C, 7 days), and DA (1°C, 0.5 m/s, 85% relative humidity [RH], 30 days).Results: Dry-aged beef (DAB) exhibited higher pH, water holding capacity (WHC), myofibrillar fragmentation index (MFI), and digestibility, however lower lightness, redness, and yellowness values, cooking loss, and shear force (SF), than those of wet-aged beef (WAB) (p<0.05). The myosin light chain band intensity of DAB was higher than that of control and WAB in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The in vitro digestibility of aged beef was highly (p<0.001) correlated to physicochemical properties except WHC. The correlation coefficient between AMs and WHC was higher than that between AM and SF (p<0.05) or MFI (p<0.001). A high correlation was observed between SF and MFI (p<0.001).Conclusion: Thus, we believe that DAB is more advantageous than WAB owing to its high digestibility and WHC and low SF.

scholarly journals Regulation of Sialic Acid Catabolism by the DNA Binding Protein NanR in Escherichia coli

2003 ◽  
Vol 185 (16) ◽  
pp. 4806-4815 ◽  
Author(s):  
Kathryn A. Kalivoda ◽  
Susan M. Steenbergen ◽  
Eric R. Vimr ◽  
Jacqueline Plumbridge
Keyword(s):  
Escherichia Coli ◽  
Sialic Acid ◽  
Promoter Region ◽  
Tandem Repeats ◽  
Polyacrylamide Gel Electrophoresis ◽  
Microbial Pathogens ◽  
Fusion Strain ◽  
Exogenous Addition

ABSTRACT All Escherichia coli strains so far examined possess a chromosomally encoded nanATEK-yhcH operon for the catabolism of sialic acids. These unique nine-carbon sugars are synthesized primarily by higher eukaryotes and can be used as carbon, nitrogen, and energy sources by a variety of microbial pathogens or commensals. The gene nanR, located immediately upstream of the operon, encodes a protein of the FadR/GntR family that represses nan expression in trans. S1 analysis identified the nan transcriptional start, and DNA footprint analysis showed that NanR binds to a region of ∼30 bp covering the promoter region. Native (nondenaturing) polyacrylamide gel electrophoresis, mass spectrometry, and chemical cross-linking indicated that NanR forms homodimers in solution. The region protected by NanR contains three tandem repeats of the hexameric sequence GGTATA. Gel shift analysis with purified hexahistidine-tagged or native NanR detected three retarded complexes, suggesting that NanR binds sequentially to the three repeats. Artificial operators carrying different numbers of repeats formed the corresponding number of complexes. Among the sugars tested that were predicted to be products of the nan-encoded system, only the exogenous addition of sialic acid resulted in the dramatic induction of a chromosomal nanA-lacZ fusion or displaced NanR from its operator in vitro. Titration of NanR by the nan promoter region or artificial operators carrying different numbers of the GGTATA repeat on plasmids in this fusion strain supported the binding of the regulator to target DNA in vivo. Together, the results indicate that GGTATA is important for NanR binding, but the precise mechanism remains to be determined.

scholarly journals Molecular basis for heterogeneity of the human p53 protein.

1986 ◽  
Vol 6 (12) ◽  
pp. 4650-4656 ◽  
Author(s):  
N Harris ◽  
E Brill ◽  
O Shohat ◽  
M Prokocimer ◽  
D Wolf ◽  
...  
Keyword(s):  
Electrophoretic Mobility ◽  
P53 Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Antigenic Determinants ◽  
Sequence Difference ◽  
Cdna Clones ◽  
Protein Species ◽  
Free System ◽  
The Individual

The human p53 tumor antigen comprises several physically distinct proteins. Two p53 proteins, separable by polyacrylamide gel electrophoresis, are expressed by the human transformed cell line SV-80. The individual cDNAs which code for these proteins were isolated and constructed into the SP6 transcription vector. The proteins encoded by these clones were identified by in vitro transcription with the SP6 vector and translation in a cell-free system. p53-H-1 and p53-H-19 cDNA clones code for the faster- and slower-migrating p53 protein species, respectively, of SV-80. The in vitro-expressed proteins of p53-H-1 and p53-H-19 had the same antigenic determinants and were structurally indistinguishable from their in vivo counterparts. By expressing defined restricted cDNA fragments in vitro, the region of heterogeneity between the respective cDNAs was located at the 5' end of the cDNAs. Exchanging the 5' fragments of interest and expressing the chimeric clones in vitro confirmed that the DNA heterogeneity was responsible for the difference in the electrophoretic mobility of these proteins. The sequences of the two cDNAs revealed a single base pair difference (G versus C) in the coding region of the clones. This sequence difference resulted in an arginine being coded for in clone p53-H-1 and a proline being coded for at the equivalent position in clone p53-H-19. This variation accounted for the change in the electrophoretic mobility of the individual p53 protein species.

Role of Surface Negative Charge in Platelet Function Related to the Hyperreactive State in Estrogen-Treated Prostatic Carcinoma

1982 ◽  
Vol 47 (03) ◽  
pp. 203-209 ◽  
Author(s):  
Stephanie M Jung ◽  
Kenji Kinoshita ◽  
Kenjiro Tanoue ◽  
Ichiro Isohisa ◽  
Hiroh Yamazaki
Keyword(s):  
Sialic Acid ◽  
Platelet Function ◽  
Prostatic Cancer ◽  
Negative Charge ◽  
Population Changes ◽  
Platelet Surface ◽  
The Relationship ◽  
Dose Dependent

SummaryThe relationship between platelet surface negative charge and hyperfunction was examined by determining electrophoretic mobility (EPM), aggregability, and sialic acid of platelets in prostatic cancer, prostatic cancer with estrogen, prostatic cancer with estrogen and aspirin, prostatic hypertrophy, and healthy aged males. Estrogen treated prostatic cancer patients had significantly higher platelet EPM. A good linear correlation was found between sialic acid and EPM (r = 0.97, p <0.001). EPM was negatively correlated with primary aggregations by adrenaline and ADP but not with secondary or maximum aggregations, suggesting increased surface negative charge may inhibit primary aggregation. Estrogen and platelet population changes influenced surface negative charge. Neuraminidase removal of platelet surface sialic acid resulted in dose-dependent decreases of EPM which paralleled decreases in sialic acid. Aspirin treated patients and platelets incubated with aspirin in vitro both showed increased platelet EPM. These results suggest that platelet surface negative charge may directly affect platelet function.

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