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J Järnefelt

Publications and source records attributed to J Järnefelt.

18 recordsLinked to original sources

Diffusion of fluorescein-labelled molecules in suspensions of erythrocyte ghosts.

The diffusion of fluorescein isothiocyanate-labelled dextran molecules in suspensions of centrifugally, tightly packed, erythrocyte ghosts was measured by fluorescence recovery after photobleaching. In comparison with diffusion in aqueous solution, the diffusion coefficients for probe molecules of varying size were about two orders of magnitude smaller. It was established that the dextran molecules remained in the space between the ghosts. Since crosslinking membrane surface carbohydrates with antibodies further inhibits diffusion, it is assumed that interactions between surface carbohydrates and the probe molecules are the cause of slow diffusion. Two alternative models are discussed.

Antibodies↗

Glycosylation of proteins in developing human brain.

The carbohydrate content of human brain glycoproteins was studied during development from the age of 12 fetal weeks to 8 postnatal months. The concentration of all the glycoprotein monosaccharides increased with age. The total amount of glycoprotein monosaccharides per lipid-free dry tissue increased by about 150% from the end of the third fetal month to the time of delivery. The increase leveled off around term, and only minor increase occurred after birth. The adult level was reached by the fifth postnatal month.

Adult↗

Blood-group A and B determinants are located in different polyglycosyl peptides isolated from human erythrocytes of blood-group AB.

The distribution of blood-group A and B determinants was studied by isolating blood-group ABH-active polyglycosyl peptides from delipidated human blood-group AB erythrocyte membranes after extensive digestion with pronase followed by chromatography on Bandeiraea simplicifolia I (BsI) lectin coupled to Sepharose. 20% of the polyglycosyl peptides were bound to BsI lectin. The glycopeptides bound were further fractionated using the blood-group-A-specific lectin from Vicia cracca (Vc). Approximately half of these were bound to the Vc lectin. The glycopeptides, which were bound to the Vc column, were not bound to the blood-group-B-specific isolectin from B. simplicifolia (BsIB4) whereas the Vc-unbound glycopeptides readily bound. The results indicate that in the polyglycosyl peptides isolated from AB erythrocytes A and B determinants are located in different carbohydrate chains. The polyglycosyl peptides, which did not bind to BsI lectin, were composed on the average of 30 monosaccharide units and those that bound contained on the average 55 monosaccharide units. The sugar composition was similar in both fractions except that N-acetylgalactosamine was found only in the BsI-bound glycopeptides. The substitution patterns of the monosaccharides were quite similar in both fractions except 2,3-O-linked galactose, which was enriched 7.5-fold in the BsI-bound glycopeptides and 3,6-O-linked galactose, which also enriched in the BsI-bound glycopeptides suggesting that these have a more branched structure than the BsI-unbound glycopeptides. Glycopeptides derived from bands 3 and 4.5 were prepared from A1B-blood-group erythrocyte membranes and fractionated as above. 25% of the glycopeptides were bound to BsI-lectin from both samples. 70% of the BsI-bound material from band 3 was bound to Vc lectin and 60% from band 4.5. The results indicate heterogeneity in the glycosylation of these bands.

ABO Blood-Group System↗

Molecular nature of the blood-group ABH antigens of the human erythrocyte membrane.

The oligosaccharide structures specifying the blood-group ABH determinants occur in the human erythrocyte membrane in different classes of compounds. The majority occur in a novel class of complex carbohydrate chains called the polyglycosyl chains. They are bound by an alkali-stable bond to glycoproteins (band 3, band 4.5) and occur also in glycolipids. Conventional glycosphingolipids as well as alkali-labile carbohydrate chains of glycoproteins (in the PAS-bands) are also carriers of the blood-group determinants.

ABO Blood-Group System↗

Enhancement of the Yu and Ledeen gas-liquid chromatographic method for sialic acid estimation: use of methane chemical ionization mass fragmentography.

The sialic acid present in erythrocyte ghosts was estimated by methane chemical ionization mass fragmentography of the trimethylsilyl methyl glycosides. The internal standard was 3,4,6-tris-trimethylsilyl-alpha-phenyl-2-deoxy-2-acetamide-D-glucosaminide, as proposed by Yu and Ledeen (J. Lipid Res. 1970. 11: 506-516). The [MH-16]+ions (m/e 610 for the TMS-methylglycoside of N-acetylneuraminic acid and m/e 498 for the internal standard) were used for quantifying nanogram levels of sialic acid in the presence of other contaminating substances. This effectively raises the signal-to-noise ratio for the Yu and Ledeen method by at least two orders of magnitude. The sensitivity and linearity of the method, without use of isotopic carriers, were tested using known quantities of N-acetylneuraminic acid. The limit of detection was below 0.4 nanograms (approximately one picomole). The useful range of detection was 10 ng-1 microgram, showing a large dynamic range.

Erythrocyte Membrane↗

Erythroglycan, a high molecular weight glycopeptide with the repeating structure [galactosyl-(1 leads to 4)-2-deoxy-2-acetamido-glucosyl(1 leads to 3)] comprising more than one-third of the protein-bound carbohydrate of human erythrocyte stroma.

Glycopeptides of molecualr weight range 7,000 to 11,000, unusual in size and structure, have been partially purified from pronase digests of lipid-free human erythrocyte ghosts; we term this fraction "erythroglycan." These substances comprise about one-third of the galactose and glucosamine of the ghost. Methylation analysis of erythroglycan yields mainly 4-linked glucosamine, 3-linked galactose, and 3,6-linked galactose, along with mannose and fucose derivatives. Hydrazinolysis and nitrous acid deamination degrade erythroglycan to galactosyl-2,5-anhydromannose, indicating a repeating structure of galactosyl, (1 leads to 4)-2deoxy-2-acetamidoglucosyl (1 leads to 3). Digestion with the endo-beta-galactosidase from Escherichia freundii gives only partial cleavage of the erythroglycan, probably because of the arborized structure indicated by the branched galactose. Since sphingosine is not detectable after methanolysis by chemical ionization mass spectrometry, and since amino acids are present, we conclude that these substances are probably glycopeptidic in origin and are not "macroglycolipids." Erythroglycan may have the same type of keratan-like core structure as the long chain blood group glycolipids from human erythrocytes and could be a protein-bound carrier of the ABO determinants.

Anion Exchange Protein 1, Erythrocyte↗

Factors affecting the relative magnitudes of the ouabain-sensitive and the ouabain-insensitive fluxes of thallium ion in erythrocytes.

A maximal rate of the ouabain-sensitive 204-Tl influx in human erythrocytes can be attained at trace concentrations of Tl+ in Mg2+ isotonic media free of K+ and Na+. The maximal influx of Tl+ from isotonic Mg(NO3)2 at 20 degrees C and pH 7.4 was 0.45 mM.l(-1).h-1 with a Km of 0.025 mM. In contrast to the active influx of Tl+, the passive Tl+ fluxes were neither saturated nor influenced by external cations in the range of concentrations of Tl+ and K+ studied. The rate constants of Tl+ passive fluxes in human and cat erythrocytes can be related to pH by the equation log kin(OUT)= -A + B.pH, where A and B are empirical constants for particular conditions. The apparent activation energy was 16 and 11 kcal/mol in sulphate and nitrate media, respectively. Tl+ and the alkali metal cations seem to overcome a common barrier in the erythrocyte membrane. Nevertheless, the rate of the passive penetration of Tl+ is about two orders of magnitude faster than those of K+ or Rb+. An extra non-Coulombic interaction between Tl+ and membrane ligands appears to be involved providing an accumulation of Tl+ somewhere in the vicinity of the membrane barrier and increasing the diffusion fluxes of Tl+ in both directions.

Animals↗

Protein-lipid interactions in the sialic acid incorporating system of liver microsomes.

The conditions for the incorporation of sialic acid (N-acetylneuraminic acid) from CMP-sialic acid into endogenous acceptors of rat liver microsomes has been studied. It is shown that the incorporating activity can be solubilized by extraction of the microsomes with a mild detergent, Triton X-100. The specific activity of the soluble system is about sixfold compared to the original microsomes. Removal of lipids from the system greatly reduces its ability to incorporate sialic acid. Recombination with phospholipids prepared from liver microsomes restores the activity. Other lipids are ineffective, and single phospholipid fractions are less effective than the phospholipid mixture. It is concluded that the system studied, comprising both sialyl transferase and sialyl acceptor-protein is a typical intrinsic membrane protein system, dependent on a hydrophobic environment for full activity.

Acetone↗

Thallium inhibition of ouabain-sensitive sodium transport and of the (Na+ plus K+)-ATPase in human erythrocytes.

The influence of Tl+ on Na+ transport and on the ATPase activity in human erythrocytes was studied. 0.1-1.0 mM Tl+ added to a K+-free medium inhibited the ouabain-sensitive self-exchange of Na+ and activated both the ouabain-sensitive 22Na outward transport and the transport related ATPase. 5-10mM external Tl+ caused inhibition of the ouabain-sensitive 22Na efflux as well as the (Na+ plus Tl+)-ATPase. Competition between the internal Na+ and rapidly penetrating thallous ions at the inner Na+-specific binding sites of the erythrocyte membrane could account for the inhibitory effect of Tl+. An increase of the internal Na+ concentration in erythrocytes or in ghosts protected the system against the inhibitory effect of high concentration of Tl+. A protective effect of Na+ was also demonstrated on the (Na+ plus Tl+)-ATPase of fragmented erythrocyte membranes studied at various Na+ and Tl+ concentrations.

Adenosine Triphosphatases↗