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R Brossmer

Publications and source records attributed to R Brossmer.

At least 91 records · Page 5Linked to original sources
PubMed1984

Determination of methylumbelliferyl-N-acetylneuraminic acid sialidase for clinical purposes.

In order to determine optimal conditions for the clinical application of leukocyte acid sialidase determination, we have studied some characteristics of the enzyme giving attention more specifically to the enzyme's thermolability and the influence of potential effectors. The enzyme has a pH optimum around 4 and a Km value of 0.25 mmol . 1(-1). A decrease of activity is observed when the enzyme is measured in leukocytes isolated from total blood or leukocytes containing plasma kept for several hours at room temperature. On the other hand, in a leukocyte homogenate, the enzyme is stable at 0 degree C for 8 hours although pronounced thermolability is observed at 23 and 37 degrees C and total inactivation occurs after 10 minutes at 50 degrees C. The addition of albumin to the homogenate partially protects the enzyme. Residual activities of 40% and 50% were measured in leukocytes and leukocyte homogenates frozen during 2 months at -20 degrees C. A marked inhibition was obtained by Triton X-100, sodium taurocholate, and methoxyphenyl-N-acetylneuraminic acid. Immediate isolation of the leukocytes after the blood drawing and subsequent homogenization at 0 degree C of a dense pellet make it possible to achieve enzyme linearity and reproducible results.

Colorimetry↗
PubMed1983

On the specificity of sialidase. Synthesis and properties of N5-acetyl-beta-D-neuraminoylpeptides - AcNeu-Gly-OH, AcNeu-Glu-OH, AcNeu-Phe-OH - and the corresponding alpha-ketosides.

By means of the mixed anhydride procedure the benzyl alpha-ketoside of N5-acetyl-D-neuraminic acid was linked to L-glycine, L-glutamic acid and L-phenylalanine. Hydrogenolytic cleavage of the benzyl group resulted in the corresponding free N5-acetyl-beta-D-neuraminoylpeptides. This new class of compounds is no substrate for Vibrio cholerae sialidase. The enzyme does not split the benzyl alpha-ketosides of N5-acetyl-D-neuraminoylpeptides nor is its activity inhibited by these compounds. The results strongly support the assumption that in sialidase substrates the carboxy group must be located close to the ketosidic oxygen. N-(N5-acetyl-beta-D-neuraminoyl)-L-phenylalanine was readily hydrolysed by carboxypeptidase A from bovine pancreas.

Animals↗
PubMed1981

Effects of chemical modification on the conformation and biological activity of peanut agglutinin.

The effect of chemical modifications on the biological properties of peanut agglutinin was investigated. The free amino groups were modified with succinic anhydride and 1-isothiocyanato-4-benzenesulfonic acid. Though the extent of modification was 95 and 85%, respectively, these derivatives did not lose their sugar binding capacity. The agglutinating activity with neuraminidase-treated human erythrocytes and various tumor cells was reduced. The mitogenic activity tested with neuraminidase-treated human lymphocytes was also diminished The tyrosine residues were modified with tetranitromethane and further with 4-aminophenyl-alpha-D-glucopyranoside and the negatively charged 2-(4-amino-benzyl)-alpha-D-neuraminic acid. The extent of modification was 30, 28 and 6%, respectively. The agglutinating and mitogenic activities were in this case not severely changed. The influence of all these modifications on the conformation was investigated by means of CD studies in the far and near ultraviolet regions.

Agglutination↗
PubMed1981

[1H-NMR spectroscopy--a potent method for the determination of substrate specificity of sialidases (author's transl)].

We describe here the application of 1H-NMR spectroscopy to determine the substrate specificity of sialidases using a 1:1 mixture of NeuAc alpha 2-3Gal beta 1-4Glc and NeuAc alpha 2-6Gal beta 1-4Glc, one viral and five bacterial sialidases. This method utilizes the separate signals in NMR spectra, characteristic for the different alpha ketosidically linked NeuAc residues and also for bound and free NeuAc. The signals generally most suitable for these purposes are those of H3a, H3e and NCOCH3. By observation and integration of these signals we can follow--qualitatively and quantitatively--which and how many NeuAc residues of the substrates are hydrolized. In contrast to the generally used colorimetric tests it is now possible to investigate with this method substrates containing two or more NeuAc residues and to determine the corresponding rate constants for hydrolysis of the differently bound NeuAc molecules. The six sialidases used show large differences in their specificity as compared with our "model substrate": The sialidase from fowl plague virus hydrolizes NeuAc alpha 2-3Gal beta 1-4Glc nearly 18 times and the enzyme from Clostridium perfringens four times, from Vibrio cholerae two times faster than NeuAc alpha 2-6Gal beta 1-4Glc. On the contrary, the sialidase from Arthrobacter ureafaciens hydrolizes the alpha 2-6 linkage six times faster than the alpha 2-3 linkage. The sialidases from Bifidobacterium show no obvious differences in their specificities relative to the linkage.

Actinomycetaceae↗
PubMed1980

[1H-NMR-spectroscopic evidence for the release of N-acetyl-alpha-D-neuraminic acid as the first product of neuraminidase action (author's transl)].

The 1H-NMR spectroscopy was used to study the anomeric configuration of N-acetyl-D-neuraminic acid released by the action of neuraminidase. The hydrolysis of NeuAcalpha 2 leads to 3 Gal-beta 1 leads to 4Glc (20mM) by the enzymes of Clostridium perfringens and Arthrobacter ureafaciens (50 mU, 150 mU and 800 mU, respectively) in 50mM Na/K-phosphate buffer pD 5.4 was observed by recording the spectra. On the basis of the characteristic signals of the protons at C-3 (alphaNeuAc: delta[H(3e)] = 2.72, delta[H(3a)] = 1.64; betaNeuAc: delta[H(3e)] = 2.25, delta[H(3a)] = 1.84) the product of the enzymatic cleavage was identified to be the N-acetylneuraminic acid in the alpha-anomeric form. Two hypotheses are discussed to explain how the enzymatic hydrolysis may occur and how N-acetyl-alpha-D-neuraminic acid leaves the catalytic site of the neuraminidases with retention of the C-2 configuration.

Arthrobacter↗
PubMed1979

Synthesis of anomeric ethyl ketosides of 5-N-acetyl-D-neuraminic acid.

The syntheses of anomeric ethyl ketosides of 5-N-acetyl-D-neuraminic acid are described. The alpha-anomer prepared by a modified Koenigs-Knorr procedure starting from acetochloroneuraminic acid is quantitatively cleaved by Vibrio cholerae neuraminidase. Proton-catalyzed reaction of 5-N-acetyl-D-neuraminic acid with ethanol yields the beta-anomer.

Chemical Phenomena↗
PubMed1978

Inhibition of human anti-Gd cold agglutinins by sialyllactose.

Two examples of IgM kappa-monotopic cold agglutinins occurring in patients with non-Hodgkin lymphomas, reacted with erythrocyte autoantigens, which were protease-resistant but were inactivated by neuraminidase. The cold agglutinins were inhibited by the trisaccharide sialyllactose [NeuNAc(alpha,2 leads to 3) Gal (beta, 1 leads to 4)Glc], which is not related to oligosaccharides known to inhibit anti-I/-i cold agglutinins, Anti-Pr cold agglutinins are not inhibited by sialyllactose, although N-acetylneuraminic acid (NeuNAc) is an essential component not only of Gd but also of Pr determinants. Gd determinants are not bound to erythrocyte membrane glycoproteins, but are apparently bound to membrane glycolipids (gangliosides).

Agglutinins↗
PubMed1978

The effect of amphiphilic phenylalkyl derivatives on platelet energy metabolism. Stimulation of glycolysis through activation of membrane ATPase.

In order to obtain information about the target of membrane-active inhibitors of platelet aggregation two phenylalkanoles and two phenylalkylamines were examined with respect to their influence on membrane ATPase and energy metabolism. The phenylalkanoles, 2-phenylethanol and 3-phenylpropanol, strongly enhanced the liberation in washed platelets of inorganic phosphate (Pi) from endogenous substrate or added ATP. A simultaneous decrease was found in the ATP/ADP ratio while glucose uptake, glycogen utilization and lactate formation increased. The effects of 2-phenylethanol and 3-phenylpropanol on Pi liberation and ATP/ADP ratio were detectable only with starving platelets; the stimulation of glycolysis could also be seen when glucose was added. It is concluded that presence of glucose enabled the platelets to reincorporate additionally liberated Pi into their ATP pool by virtue of the higher metabolic rate. The data suggest that a membrane-ATPase, possibly thrombosthenin ATPase, is the target of the phenylalkanoles. The phenylalkylamines, 2-phenylethylamine and 4-phenylbutylamine, appeared to inhibit Pi liberation and energy metabolism. Hence the aggregation inhibition produced by amphiphilic phenylalkylamines and phenylalkanoles is not due to a uniform metabolic effect of both classes of derivatives.

Adenosine Diphosphate↗