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Biomedical subjects

J Hase

Publications and source records attributed to J Hase.

At least 19 recordsLinked to original sources
PubMed1984

Modification of thiol groups of Jack bean urease with diazonium-1H-tetrazole.

Jack bean urease [EC 3.5.1.5] was modified with diazonium-1H-tetrazole (DHT). Reaction of DHT with the enzyme produced a characteristic absorption peak at 320 nm and led to complete loss of the enzymatic activity at a low concentration of DHT. Amino acid analysis of DHT-modified urease showed that only cysteine residues reacted with the reagent and other amino acid residues such as tyrosine, histidine, and lysine did not. The enzymatic activity was protected against DHT-inactivation by the addition of substrate. On the other hand, when the cysteine residues were modified with DHT, the enzyme was not converted to polymeric forms. Furthermore, the binding ability of urease with hydroxamic acid, a specific urease inhibitor, was virtually unaffected by DHT-inactivation. These results indicate that cysteine residues are specifically modified by DHT with concomitant loss of enzymatic activity and polymerization ability, but are not essential for the binding of hydroxamic acid to the enzyme.

Amino Acids↗
PubMed1983

Effect of fructose 1,6-bisphosphate on the activity of liver pyruvate kinase after limited proteolysis with cathepsin B.

Treatment of rat liver-type pyruvate kinase with rabbit liver cathepsin B at pH 7.0 caused loss of activity in the standard assay with 0.6 mM of phosphoenolpyruvate. The modified enzyme exhibited about 10% of the original activity when assayed with 2.0 mM of the substrate. No detectable change in the subunit molecular weight of the enzyme occurred during inactivation. On addition of 4 microM fructose 1,6-bisphosphate the activity of the treated enzyme was restored to that of the original enzyme. Limited proteolysis of the enzyme by cathepsin B appears to enhance the requirement for the positive effector, fructose 1,6-bisphosphate.

Animals↗
PubMed1983

Photo-oxidation of Jack bean urease in the presence of methylene blue.

Photo-oxidation of Jack bean urease was performed in the presence of a low concentration of methylene blue, which led to the complete loss of the enzymatic activity. The inactivation was more remarkable in an alkaline region than in an acidic region and prevented by the addition of histidine or methionine. Amino acid analysis of the oxidized enzyme revealed that the number of histidine residues had decreased to 73% that of the native enzyme, but the numbers of other amino acid residues were not significantly affected. Benzohydroxamic acid, a specific urease inhibitor, protected the active site of the enzyme against photo-oxidation. On the other hand, oxidation of the enzyme decreased its binding ability with caprylo- and benzohydroxamic acid to one-third. These results suggest that histidine residues are modified by photo-oxidation and are essential to both the enzymatic activity and the binding ability with hydroxamic acid.

Fabaceae↗
PubMed1983

Cholesterol-lowering effect of Clostridium butyricum in cholesterol-fed rats.

We examined the effects of oral administration of Clostridium butyricum miyairi on the catabolism and excretion of cholesterol in normal rats by using 14C-labeled cholesterol, and on plasma cholesterol levels in the rats and mice fed a high cholesterol diet. This bacterial treatment (about 10(7) cells/day) accelerated the cleavage of cholesterol side chain in normal rats. Plasma cholesterol levels of rats fed the high cholesterol diet with the bacterium (about 10(9) cells/day) were significantly lower than those of the control rats, and the effect was confirmed by exchanging the diets of both groups. However, plasma cholesterol levels of mice were not affected by this bacterial treatment.

Administration, Oral↗
PubMed1983

Study on hydroxamic acids and their urease inhibitory potency by quantum chemistry calculation.

The electronic structures of 34 hydroxamic acids [R-(CONHCH2(n-CONHOH, R = aromatic or aliphatic, n = 1 or 0] were calculated by the INDO method and their urease inhibitory potencies were discussed in terms of the calculated electronic parameters and molar refraction. The charge distribution of -CONHOH residue which has been expected as a functional group for inhibition slightly be affected by the change of R - moiety and by the presence or absence of the -CONHCH2- residue. The best improved regression equation indicated that the inhibitory potency of hydroxamic acids was parabolically varied with the molar refraction and that the increase of the inhibitory potency by the presence of -CONHCH2- residue was explained by the variation of the charge density of a carbon atom directly bonding the -CONHOH group.

Chemical Phenomena↗
PubMed1982

Effects of cholesterol evulsion on susceptibility to perfringolysin O of human erythrocytes.

Human erythrocytes preincubated with a phosphatidylcholine suspension (preincubated cells) showed decreased susceptibility to perfringolysin O, the decrease being strongly affected by preincubation time and temperature, and the phosphatidyl choline concentration. The binding of the toxin to the preincubated cells also decreased with the preincubation time and reached minimum at 37 degrees C for 6 h. Through this preincubation, about 30% of cholesterol was removed from cells without lysis. The susceptibility of preincubated cells to the toxin seemed to be affected by the amount of cholesterol removed from cells, but not by the cholesterol content of cell membranes. This indicates that most of the cholesterol interactive with the toxin is removable from cell membranes by preincubation with phosphatidylcholine suspension, and that the residual cholesterol is firmly constituted in the membrane structure and cannot interact with the toxin. After cholesterol evulsion by the preincubated plasma method (Murphy, J.R. (1962) J. Lab. Clin. Med. 60, 86-109 and 60, 571-578), cells also exhibited lower susceptibility to the toxin and to saponins, but higher susceptibility to lysophosphatidylcholine.

Bacterial Toxins↗
PubMed1982

Age-development changes in susceptibility of erythrocytes to perfringolysin O.

Susceptibility to perfringolysin O of erythrocytes from mice of different ages was examined. Erythrocytes of mice younger than 5 weeks' old were more resistant to the toxin than those of young adult and adult mice. Erythrocytes of aged mice were about 3.5 times more susceptible to the toxin than erythrocytes from 4-week-old mice. The membrane cholesterol content of erythrocytes appeared to be maintained at a constant level throughout the ages of mice examined. About 5% of the total membrane cholesterol was supposed to provide receptor sites for the toxin from an experiment in which cholesterol was specifically extracted by liposomes. It was demonstrated in this experiment that susceptibility of erythrocytes to the toxin was lost in proportion to the reduction in the toxin binding. The susceptibility, however, of erythrocytes from young or aged mice was much lower or higher than expected from the changes in toxin binding. Therefore, two possibilities were raised to account for age-related alterations in the susceptibility of erythrocytes; not only expansion of a particular compartment of membrane cholesterol as a toxin receptor but also some activation of intracellular reactions leading to hemolysis might occur in senescence.

Aging↗
PubMed1982

High-molecular-weight hemolysin of Clostridium tetani.

Clostridium tetani excretes hemolysins of two size classes, a high-molecular-weight hemolysin (HMH), which was eluted near void volume of a Sepharose 6B column, and conventional tetanolysin (molecular weight, approximately 50,000). The total hemolysin activity in the culture supernatant increased sharply with growth of bacteria and remained at a high level during autolysis. The content of HMH, however, decreased from 41% at 4 h of culture to 0.4% at the early stage of autolysis. The cell bodies also exhibited hemolytic activity, 70% of which could be solubilized and separated into HMH and the 50,000 Mr tetanolysin as extracellular hemolysins. The activity ratio of HMH to the total solubilized hemolysins was 0.45, on the average, at 6 h of culture but was 0.23 at the middle of logarithmic growth. Partially purified HMH from both sources appeared as broken pieces of cytoplasmic membranes under an electron microscope. The ratio of proteins to phospholipids in HMH was found to 3.26, a value similar to that in cell membrane. The total cell hemolytic activity decreased by 90 or 75% upon addition of chloramphenicol or anti-tetanolysin serum, respectively, into a 6-h-old culture of bacteria. It is suggested that HMH is a complex of tetanolysin with a membrane fragment and releases the conventional tetanolysin during bacterial culture.

Bacterial Toxins↗
PubMed1981

The structure-hemolysis relationship of oleanolic acid derivatives and inhibition of the saponin-induced hemolysis with sapogenins.

Chikusetsusaponin IV and V, whose genin is oleanolic acid, exhibited weak hemolytic activities. Removal of glucose residue at position 29 of chikusetsusaponin V by partial hydrolysis increased the activity more than 30-fold. Methylation of the carboxyl group at position 28 increased the activity furthermore by about 10-fold, showing HD50 value of 3.77 microM. On the other hand, removal of the sugar chain at position 3 of chickusetsusaponin V by partial hydrolysis completely lost the activity. These facts suggest that the sugar chain at position 3 of oleanolic acid is essential but that at position 29 is pernicious for the activity. The cytolytic agents, whose target has been regarded as membrane cholesterol, were inactivated not only by cholesterol but also by sapogenins such as oleanolic acid, gitogenin and hederagenin. Among saponins tested, akebia saponin B and C were inactivated by cholesterol, but not by the genins, probably because their affinities for the genins are too low to form complexes.

Anti-Bacterial Agents↗
PubMed1980

Purification and some properties of tetanolysin.

Tetanolysin was purified from the culture fluid of a strain of Clostridium tetani by ammonium sulfate fractionation, acetone precipitation and repeated gel filtration. Two hemolysins with different molecular weights were separated by gel filtration, and the smaller one, tetanolysin, was further purified. The purification raised the specific activity of tetanolysin 1,050-fold to 500 HU/micrograms of protein. The purified preparation gave a single, relatively broad band on polyacrylamide gel electrophoresis, in which the activity was roughly parallel with the protein concentration. However, on sodium dodecylsulfate-gel electrophoresis it gave two bands with nearly equal amounts of proteins, showing molecular weights of 53,000 and 48,000 +/- 3,000. Furthermore, isoelectric focusing revealed four peaks of the activity whose isoelectric pHs were 6.1, 5.6, 5.3, and 6.6 in decreasing order of the activity. These findings suggest that the preparation contains four hemolysins with different pIs, which are classifiable into two groups by molecular size. The preparation was completely free of tetanus neurotoxin and proteases. Tetanolysin was more strongly inhibited by cholesterol and more rapidly absorbed onto erythrocytes than theta-toxin of Cl. perfringens.

Cholesterol↗
PubMed1980

Therapy for urolithiasis by hydroxamic acids. II. Urease inhibitory potency and urinary excretion rate of hippurohydroxamic acid derivatives.

The apparent I50 values of various hippurohydroxamic acids against urease activity of sword bean were mostly 0.5 to 2.0 microM regardless of hydrophobicity of their substituents. However, the marked increase of hydrophilicity caused by substitution of trimethoxy groups conspicuously decreased the inhibitory potency. Methylation at alpha-position of the hydroxamic acid group in these compounds remarkably decreased the inhibitory potency, probably owing to steric hindrance by the alpha-methyl group. Thenoyl-, furoyl- and nicotino-glycinohydroxamic acids which are bioisostereomers of hippurohydroxamic acid had I50 values of 0.64, 1.3 and 5.3 microM, respectively. Furthermore, the inhibitory potency of some substituted hippurohydroxamic acids against the ureolytic activity of intact Proteus mirabilis isolated from patients with urinary tract infection, were half to one-tenth of those against urease activity of sword bean. On the other hand, m- and p-nitro-, m- and p-methoxy-, m- and p-acetylamino-hippurohydroxamic acid and furoylglycinohydroxamic acid showed high urinary excretion rates of 14 to 16% of the doses administered orally to rats, while most of the others had excretion rates of about 3 to 5%.

Fabaceae↗
PubMed1980

Therapy for urolithiasis by hydroxamic acids. III. Urease inhibitory potency and urinary excretion rate of N-acylglycinohydroxamic acids.

Hydroxamic acid, a potent urease inhibitor, having a high urinary excretion rate is expected to be a therapeutic agent for urolithiasis caused by urea-splitting bacterial infection of the urinary tract. Twenty-one new derivatives of N-aliphatic-acylglycinohydroxamic acids (GHAs) were synthesized, and their inhibitory potencies against the urease activity of sword bean in a phosphate buffer and against the ureolytic activity of Proteus mirabilis in human urine, and their urinary excretion rates in rats were also measured for this purpose I50 values of most of GHAs against the urease activity of sword bean were about 1 to 10 microM and 2-ethyl-n-butyroyl GHA was the most potent inhibitor with the value of 0.79 microM. I50 values of most of the GHAs against the ureolytic activity of Proteus mirabilis were about 5 to 50 microM and n-nonaroyl GHA was the most potent inhibitor with the value of 3.6 microM. 2,2-Dimethylpropionyl GHA had the highest urinary excretion rate with the recovery of 11%. Routes of administration of 2,2-dimethylpropionyl GHA and sex of rats used did not affect the amount of urinary excretion at all. The results in this report suggest that DL 2-methyl-n-butyroyl, 2-ethyl-n-butyroyl and 2,2-dimethylpropionyl GHA are the most hopeful therapeutic agents for urolithiasis among them.

Animals↗
PubMed1980

Quantitative structure-activity relationships between hydroxamic acids and their urease inhibitory potency.

Quantitative structure activity relationships between physico-chemical properties of four series of more than sixty hydroxamic acids [R-(CONHCH2)n-CONHOH, R=aromatic or aliphatic, n=1 or 0) and their urease inhibitory activities were examined. The best improved regression equation primarily indicated that the inhibitory activities of hydroxamic acids were parabolically varied with the hydrophobic variable, pi of the R moiety and the optimal pi value was calculated to be 2.58. Furthermore, the inhibitory activities of congeners were found to be significantly affected by the B1 variable representing the minimum width of steric size of the R moiety. Besides, it was clarified by using two indicator variables that inhibitory activities were not affected by the R moiety, whether aromatic or aliphatic, but positively affected by the presence of the -CONHCH2-group between the R moiety and the hydroxamic acid group.

Chemical Phenomena↗
PubMed1979

Alteration of human erythrocyte plasma membranes by perfringolysin O as revealed by freeze-fracture electron microscopy. Studies on Clostridium perfringens exotoxins V.

When human erythrocyte membranes were treated with perfringolysin O (Clostridium perfringens theta-toxin) and examined by electron microscopy after freeze-fracture, two ultrastructural alterations were observed in fracture faces of membrane. (1) A random aggregation of intramembranous particles was seen in the fracture face of the protoplasmic half (PF face) of all membranes treated with the toxin, even if at a low concentration (40 hemolytic units/ml). On the other hand, the aggregation in the fracture face of the exoplasmic half (EF face) was observed only in membranes treated with a high concentration (3300 hemolytic units/ml) for 2 h. (2) Round protrusions and "cavities" with 30 nm in diameter were visible in EF and PF faces of membranes treated with a high concentration, respectively. These structures were always protruded toward cytoplasmic side, but did not appear to form holes through the membrane. Ring and arc shaped structures with a dark center of 26 nm and a distinct border of 5 nm in width were observed when the toxin alone was negatively stained at a very high concentration (170,000 hemolytic units/ml). These structures were also produced in the presence of cholesterol even if the toxin concentration was low.

Bacterial Toxins↗