Heat stability and pH activity data of alpha-L-fucosidase in human serum vary with enzyme concentration

alpha-L-Fucosidase from serum of humans with either high or low enzyme activity was separately purified. the enzyme from either source had virtually the same heat stability and pH activity profile. It has been widely reported that alpha-L-fucosidase in crude sera from individuals with high and low enzyme activity differed with respect to heat stability and activity at pH 4 relative to activity at pH 5, the pH optimum of the enzyme. We investigated this discrepancy and found that both the heat stability and relative activity at pH 4 of alpha-L-fucosidase from sera with either high or low enzyme activity was dependent upon enzyme concentration. With decreasing enzyme concentration, the enzyme was more heat labile and had less relative activity at pH 4. Consequently, if the data obtained using high and low enzyme activity sera are compared on the basis of equivalent amounts of serum instead of equivalent amounts of enzyme activity, differences between the enzyme from high and low activity serum would be erroneously inferred. Apparently, this is what other investigators have done. Moreover, we found that alpha-L-fucosidase can exist in heat-stable or labile species with sedimentation coefficients of 9.8 S and 4.8 S, respectively. The interconversion and relative proportion of these species is dependent upon enzyme concentration and pH.     

宫川蜜柑根际土壤酶活性与土壤养分含量相关性的研究

研究了不同肥力水平的宫川蜜柑根际土壤酶的活性及其与土壤农化特性的关系。结果表明 :高产园的土壤酶活性显著高于低产园的土壤酶活性。经统计分析 ,土壤酶活性与养分含量均呈极显著相关。而且酶的活性在土壤中的分布有一定的规律性。其水平分布是在树冠内半径的 4 /5处至树冠滴水线范围内 ,酶的活性最高 ,由此处向内向外酶的活性逐渐降低 ;其垂直分布是 0～ 2 0 cm土层酶的活性最高 ,随土层的加深而逐渐降低     

Insulin and fructose regulate malic enzyme activity by different processes

A comparison of the regulatory processes controlling hepatic malic enzyme activity following treatment of diabetic rats with insulin or with a high fructose diet demonstrated several important differences. Insulin treatment caused a 50-fold increase in activity, due to a 12-fold increase in enzyme quantity and a 4-fold increase in specific activity(units/nmol). Dietary fructose caused a 3-fold increase in enzyme activity, due to a 3-fold increase in enzyme quantity, with no change in the specific activity of the enzyme. Thus, while fructose initiated a minor increase in malic enzyme activity, insulin was more effective, causing a substantially greater increase in enzyme activity and activating a hormone specific alteration in the catalytic activity of each enzyme molecule.     

A dual mechanism regulates the insulin stimulation of hepatic malic enzyme

The activity of malic enzyme, an important hepatic lipogenic enzyme, is stimulated in diabetic rats by insulin administration. This process was shown to involve increases in both enzyme quantity and the specific activity (units activity/nmol enzyme) of the enzyme. Therefore, the coupling of these two regulatory mechanisms was responsible for the insulin-mediated increase in malic enzyme activity.     

Distribution of post-proline cleaving enzyme in human brain and the peripheral tissues

Summary We have studied the distribution of post-propline cleaving enzyme activity in the various tissues in humans using 7-(succinyl-Gly-Pro)-4-methylcoumarinamide as substrate. The post-propline cleaving enzyme activity was high in muscle, testes, kidney and submandibular gland, but was low in the heart, mesenterium and aorta. In the brain, relatively high post-propline cleaving enzyme activity was observed in the cerebral cortex, but other brain regions showed a very low enzyme activity.On Sephadex G-100 column chromatography, enzyme activity in human kidney showed a major peak and a minor peak. The major peak coincided with the enzyme in human cerebral cortex, but was different from human serum enzyme. Diisopropylfluorophosphate, a serine protease inhibitor, strongly inhibited the enzyme activity of each active fraction. The enzyme in the cerebral cortex and kidney was inhibited by heavy metals and thiol blocking agents. However, inhibition of enzyme activity in the serum was not observed with such inhibitors. Therefore, we suppose that post-proline cleaving enzyme activity in the brain is similar, if not identical, to that in the kidney.     

N-Acetyl-Glucosaminidase Activity during Limb Regeneration in the Adult Newt

N-Acetyl-glucosaminidase activity was measured during the first 25 days of limb regeneration. It was found that the enzyme is present in the normal limb. Following amputation a significant drop was obtained at day 3. A significant increase in enzyme activity was found at day 5 followed by a second drop by day 10. For days 12–15 a second peak of enzyme activity was detected, followed by a third drop; by day 25, normal levels of enzyme activity were detected. Histochemical localization of the enzyme in tissue samples showing enzyme activity as detected biochemically (days 5 and 17 of regeneration) gave negative results. However, enzyme activity was found in the incubation medium, indicating that the enzyme is released from the cells. The peaks of enzyme activity coincide with the stages of limb regeneration where a high degree of tissue demolition and cell lysis occurs. The latter are important events in the regeneration process, cell dedifferentiation, and blastema formation.     

银杏园土壤酶活性与土壤肥力的关系研究

研究了银杏土壤酶活性与土壤肥力的关系,试图探索利用土壤酶活性作为评价银杏果园土壤肥力的指标。结果表明,有３种酶的活性与土壤肥力存在着极为密切的关系,酶活性随土壤肥力的高低而有规律地变化。根据相关分析,该土壤的脲酶、蛋白酶、转化酶的活性均与土壤有机质、全氮、全磷、碱解氮、速效磷、速效钾及银杏果实产量呈极显著相关。不同土壤酶活性之间也存在极显著的相关性。因此,土壤酶活性作为评价银杏土壤肥力水平的指标是可行的。     

Comparative studies on the leukotriene D4-metabolizing enzyme of different types of leukocytes

1. The leukotriene (LT) D4-metabolizing enzyme which catalyzes the conversion of LTD4 to LTE4, was investigated in various types of leukocytes from guinea pigs and humans. 2. In guinea pigs, the enzyme activity was present in macrophages but was hardly present in neutrophils, lymphocytes and eosinophils. 3. In humans, neutrophils, lymphocytes and macrophages all possessed the enzyme activity. However, enzyme activity varied with cell types and macrophages showed the highest enzyme activity among the leukocytes. 4. The subcellular localization of the LTD4-metabolizing enzyme was studied and leukocytes were divided into two groups: one which has the enzyme activity exclusively on the cell surface and the other which has the activity both on the cell surface and in the granules of leukocytes. 5. The enzyme activity was remarkably inhibited by o-phenanthroline and dithiothreitol and the inactivated enzyme was considerably reactivated by Co2+ and Zn2+, suggesting that the LTD4-metabolizing enzyme of leukocytes is a metalloenzyme.     

Regulation of the activity and synthesis of malic enzyme in 3T3-L1 cells

Malic enzyme activity in differentiated 3T3-L1 cells was about 20-fold greater than activity in undifferentiated cells. A new steady-state level was achieved about 8 days after initiating differentiation of confluent cultures with a 2-day exposure to dexamethasone, isobutylmethylxanthine, and insulin. This increase in enzyme activity resulted from an increase in the mass of malic enzyme as detected by immunotitration of enzyme activity with goat antiserum directed against purified rat liver malic enzyme. Malic enzyme synthesis was undetectable in undifferentiated cells and increased to about 0.2% of soluble protein in differentiated cells, suggesting that the increase in enzyme mass was due primarily to an increase in enzyme synthesis. Thyroid hormone, a potent stimulator of malic enzyme activity in hepatocytes in culture and in liver and adipose tissue in intact animals, decreased or increased malic enzyme activity in differentiating 3T3-L1 cells by about 40% when it was removed or added to the medium, respectively. Insulin, another physiologically important regulator of malic enzyme activity in vivo, had no effect on the initial rate of accumulation of malic enzyme activity in the differentiating cells and caused a 30 to 40% decrease in the final level of enzyme activity in the fully differentiated cells. Cyclic AMP, a potent inhibitor of malic enzyme synthesis in hepatocytes in culture, inhibited this process in 3T3-L1 cells by 30%. Malic enzyme is like several other enzymes in that the large increase in its concentration which accompanies differentiation of 3T3-L1 cells is due to increased synthesis of enzyme protein. However, the hormonal modulation of malic enzyme characteristic of liver and adipose tissue in intact animals does not appear to occur in differentiated 3T3-L1 cells, suggesting that differentiated 3T3-L1 cells may not be an appropriate model system in which to study the hormonal modulation of malic enzyme that occurs in liver and adipose tissue of intact animals.     

Inhibition of 5-amino levulinic acid dehydratase activity by arsenic in excised etiolated maize leaf segments during greening

In vivo as well as in vitro supply of sodium arsenate inhibited the 5-Amino levulinic acid dehydratase (5-aminolevulinate-hydrolyase EC 4.2.1.24, ALAD) activity in excised etiolated maize leaf segments during greening. The percent inhibition of enzyme activity by arsenate (As) was reduced by the supply of KNO3, but it was increased by the glutamine and GSH. Various inhibitors, such as, chloramphenicol, cycloheximide and LA, decreased the % inhibition of enzyme activity by As. The % inhibition of enzyme activity was also reduced by in vivo supply of DTNB. The enzyme activity was reduced substantially by in vitro inclusion of LA, both in the absence and presence of As. In vitro inclusion of DTNB and GSH inhibited the enzyme activity extracted from leaf segments treated without arsenate (-As enzyme) and caused respectively no effect and stimulatory effect on arsenate treated enzyme (+As enzyme). Increasing concentration of ALA during assay increased the activity of -As enzyme and +As enzyme to different extent, but double reciprocal plots for both the enzymes were biphasic and yielded distinct S0.5 values for the two enzymes (-As enzyme, 40 micromol/L and +As enzyme, 145 micromol/L) at lower concentration range of ALA only. It is suggested that As inhibits ALAD activity in greening maize leaf segments by affecting its thiol groups and/or binding of ALA to the enzyme.     

嗜热菌Anoxybacillus sp．DL3产蛋白酶条件及其酶学性质研究

目的：对Anoxybacillussp．DL3的产蛋白酶条件及其酶学性质进行研究,为下一步进行蛋白酶基因的克隆、表达提供依据。方法：应用常规方法液体培养细菌,研究温度、pH、培养基中碳源、氮源对菌株产蛋白酶的影响,硫酸铵盐析的方法提取酶液,并采用Folin法测酶活性。用紫外分光光度计在OD680hi／1测吸光值。并对提取的蛋白酶液进行酶的最适温度、pH以及酶的热稳定性和pH稳定性研究,向酶液中添加金属离子和EDTA、PMSF,研究其对酶活性的影响。结果：在培养基初始pH是6．5,培养温度为40℃时菌株产酶酶活性最大;培养基中以乳糖为碳源,酵母膏和硫酸铵为氮源,碳源与氮源的比例为1：2时,酶活最大。酶学性质研究结果显示：该酶的最适反应温度是55℃,最适反应pH是7．0;在50℃保温20min-80min内,酶活力下降幅度较小。60℃保温60min后,仍保持约60％的酶活。70℃保温60min后,残余酶活为30％。该酶在pH为6．0～8．0范围内,相对酶活差别不是很大,下降趋势大致相同。在强碱条件下,相对酶活下降很明显。Fe2＋、Cu2＋和Hg2＋对酶活性有明显的抑制作用;Ca2＋、Mg^2＋、Mn2＋等金属离子对酶活性有明显的促进作用;乙二胺四乙酸（EDTA）和苯甲基磺酰氟（PMSF）对酶活性也有一定抑制作用。结论：Anoxybacillussp．DL3所产的蛋白酶为嗜热中性蛋白酶,此酶具有较好的热稳定性和pH耐受性,该菌株具有进一步开发、利用的价值。     

Insulin stimulation of hepatic malic enzyme activity in normal and diabetic rats controlled by different regulatory processes

A comparison of the processes controlling the increase in hepatic malic enzyme activity in insulin-treated normal and diabetic rats indicated the existence of two distinct regulatory mechanisms. Livers were removed at 12, 36, and 60 h after insulin treatment of normal and alloxan-diabetic rats, and the activity, quantity, and specific activity (units/nmol), of malic enzyme was determined. In normal rats, a significant increase in activity occurred 12 h after insulin, whereas 36 h of insulin treatment was required for diabetic rats to show an increase in enzyme activity. This suggested that the return of malic enzyme activity from the depleted levels measured in diabetic rats probably involved a different sequence of events. A malic enzyme specific radioimmunoassay confirmed this. The increase in activity in insulin-treated normal rats was due to an increase in the quantity of malic enzyme. In insulin-treated diabetic rats, the increase in activity resulted from increases in both enzyme quantity and the specific activity of the enzyme, which returned to levels observed in normal rats.     

Involvement of diminution of glutathione, produced by deficiency of methionine in the diet, in the elevation of malic enzyme level in rat liver

Rats fed on a low protein diet show an increase in the specific activity of malic enzyme and a concomitant decrease of glutathione concentration. We have studied the effect on malic enzyme activity of supplementing of low protein diet with essential amino acids. Only when methionine was excluded from the diet did the specific activity of malic enzyme increase to the same extent as found in rats fed with low protein diet. Immunoprecipitation of malic enzyme indicated that specific activity changes are the result of changes in the amounts of enzyme. Under all dietary conditions studied, the increase in malic enzyme activity is associated with a decrease in the concentration of GSH. To evaluate the possible causative role of GSH in malic enzyme induction, the specific activity of malic enzyme was measured in rats treated with BSO, an inhibitor of GSH biosynthesis. The results show that in BSO-treated rats the decrease of GSH levels is also accompanied by an increase in the activity of malic enzyme.     

Liver collagen glucosyltransferase in experimental primary liver carcinoma.

Collagen glucosyltransferase activity (EC 2.4.1.66) was quantified in experimentally-induced liver carcinoma, murine schistosomiasis mansoni-induced liver fibrosis and compared to the level of enzyme activity in control liver samples. Enzyme activity in hepatoma and fibrotic tissues were 12 and 5 times the mean level of enzyme activity in the control liver tissue respectively. The level of enzyme activity in the hepatoma tissue was two times the level of enzyme activity found in the fibrotic tissue. The findings in this study provide the basis for the highly elevated serum values of this intracellular enzyme in experimentally-induced primary hepatocellular carcinoma or in human primary hepatoma. The enzyme activity may be increased in primary liver carcinoma to compensate for an increased rate of collagen synthesis.     

DIFFERENCES IN THE REGULATION OF TYROSINE AMINOTRANSFERASE IN BRAIN AND LIVER

Abstract— l -Tyrosine:2-oxoglutarate aminotransferase (EC 2.6.1.5) activity in rat brain is not regulated in the same way as in rat liver. No diurnal rhythm in the activity of the cerebral enzyme was found in rats fed ad lib. although there was a marked diurnal variation in the activity of the hepatic enzyme. In adrenalectomized rats, hydrocortisone and glucagon induced the enzyme in liver but had no effect on the enzyme in brain. In normal rats, treatment with reserpine or exposure to cold elevated the activity of the hepatic enzyme without affecting the enzyme in brain. Thus, the tyrosine aminotransferase of brain differed from the enzyme in liver since it did not exhibit diurnal variations of activity and was not affected by hormones, drugs, or stress.     

Change in Hatching Enzyme Activity during Development of the Sea Urchin, Hemicentrotus Pulcherrimus

The time course of change in hatching enzyme activity during development of embryos of the sea urchin Hemicentrotus pulcherrimus was observed. The enzyme was present in the particulate fraction in embryos until the time of hatching and was maximal at the time of hatching. Cell fractionation studies suggested the existence of an inhibitor of the hatching enzyme. This possibility was subsequently substantiated by experiments in mixtures of fractions: the activity of hatching enzyme in the particulate fraction was inhibited by the supernatant of embryos. This inhibitory factor was heat-stable and non-dialyzable, but it was not characterized further. The activity of secreted hatching enzyme was not inhibited by this factor, suggesting that the molecular forms of hatching enzyme in embryos and in the culture supernatant are different. After hatching, the amount of increase in the hatching enzyme activity in the culture supernatant was 3.5 times the amount of decrease in enzyme activity in the embryos, suggesting that the enzyme was activated during its secretion.     

Mouse Mutant Deficient in d-Amino Acid Oxidase Activity

D-Amino acid oxidase activity in the kidney homogenates of mice of seven strains was measured to search for a mutant for this enzyme. There was a consistent sex difference in the enzyme activity in these strains: male mice showed higher levels of the enzyme activity than females. In contrast to other strains, some mice of the ddY strain did not possess enzyme activity. This trait was inheritable, and a mouse stock without enzyme activity (DAO-) was established. The allele (Dao-1c) carried by the DAO- mice was recessive and behaved as a single autosomal gene in inheritance. Heterozygous mice for this gene (Dao-1+/Dao-1c) showed nearly half the enzyme activity of the wild-type homozygotes (Dao-1+/Dao-1+), suggesting that Dao-1c is a null allele and that there is a gene dosage effect on the enzyme activity.     

Studies on oestrogen-2/4-hydroxylase activity in mammalian brain, using a radioenzymatic assay method

The activity of oestrogen-2/4-hydroxylase in brain tissue was studied using a radioenzymatic assay which controlled for non-specific formation of catechol oestrogen. The ontogeny of enzyme activity in male and female rat brain was examined between 1 and 70 days of age. Until 10 days of age, both sexes showed low enzyme activity. Between days 10 and 15 enzyme activity in the female significantly increased but was unchanged in the male. Male brain enzyme activity increased between days 15 and 20 whereas activity was unchanged in the female. Both males and females showed increases in enzyme activity between 20 and 25 days of age. Thereafter enzyme activity plateaued but at all ages between day 20 and day 70 male brain enzyme activity was significantly greater than in the female. A study of the distribution of the oestrogen-2/4-hydrolase in the sheep hypothalamus revealed that the stalk/median eminence had approx 10 times greater activity than either the anterior, medial or posterior hypothalamus. These findings are discussed in relation to sexual differentiation of the brain.     

Characterization of botryococcene synthase enzyme activity, a squalene synthase-like activity from the green microalga Botryococcus braunii, Race B

The extracellular matrix of the alga Botryococcus braunii, Race B, consists mainly of botryococcenes, which have potential as a hydrocarbon fuel. Botryococcenes are structurally similar to squalene raising the possibility of a common enzyme for the biosynthesis of both. While B. braunii squalene synthase (SS) enzyme activity has been documented, botryococcene synthase (BS) enzyme activity has not been. In the current study, an assay for BS activity has been developed and used to show that many of the assay conditions for BS enzyme activity are similar to those of SS. However, SS enzyme activity is stimulated by Tween 80 while BS enzyme activity is inhibited. Moreover, BS enzyme activity was correlated with the accumulation of botryococcenes during a B. braunii culture growth cycle, which was distinctly different from the profile of SS enzyme activity. While the current results indicate a conservation of enzymological features amongst the BS and SS enzymes, raising the possibility of one enzyme capable of catalyzing both activities, they are also consistent with these two activities arising from separate and distinct enzymes.     

Effects of dietary fatty acids on hepatic 3-hydroxy-3-methylglutaryl CoA reductase activity in hamsters on a high-glucose diet

Hepatic 3-hydroxy-3-methylglutaryl CoA reductase activity in hamsters given a fat-free high-glucose diet for 21 days was approximately 20 times higher than that in chow-fed hamsters. The increase in enzyme activity by dietary glucose was affected by saturated or unsaturated fatty acids or cholesterol added to the high-glucose diet. Ethyllinoleate or ethyloleate, added to the diet at a concentration of 5%, suppressed the increase in the enzyme activity. In contrast, addition of ethylpalmitate to the diet further stimulated the increase in the enzyme activity. Addition of 2% cholesterol to the high-glucose diet moderately suppressed, and addition of both cholesterol and ethyllinoleate completely prevented, the increase in the enzyme activity. The enzyme activity closely correlated with the incidence of formation of cholesterol gallstones but not with the liver cholesterol level. Marked increase in the enzyme activity was observed by feeding the high-glucose diet to starved hamsters for even a short period. On the third day after feeding was resumed, the enzyme activity was increased 500-fold compared to that during starvation. This increase in the enzyme activity was also reduced by dietary unsaturated fatty acid esters and stimulated by a dietary saturated fatty acid ester.