Regulatory mechanisms of UDP-glucuronic acid biosynthesis in cultured human skin fibroblasts

Kinetic parameters and regulatory properties of UDPGDH extracted from cultured human skin fibroblasts were determined and compared with those of UDPGDH from cornea and epiphysial-plate cartilage. Fibroblast enzyme showed an affinity for UDPG 7 times higher than cartilage enzyme and 42 times higher than cornea enzyme. UDP-xylose acted as a co-operative allosteric inhibitor, but under the same experimental conditions fibroblast enzyme was significantly less inhibited. These results were in agreement with the different GAG production of the cells we studied. Fibroblast UDPGDH activity was regulated by the NAD/NADH ratio and it was also affected by modifications of extracellular matrix composition. A significant increase of UDPGDH affinity for UDPG was observed after the treatment of the monolayers with Chase ABC.     

Catalytic activity and substrate specificity of the flavin-containing monooxygenase in microsomal systems: characterization of the hepatic, pulmonary and renal enzymes of the mouse, rabbit, and rat

Inhibitory antibodies against NADPH-cytochrome P-450 reductase, detergent solubilization to dissociate functional interaction between the reductase and cytochrome P-450, and selective trypsin degradation have been used to characterize flavin-containing monooxygenase activity in microsomes from different tissues and species. A comparison of assay methods is reported. The native microsome-bound flavin-containing monooxygenase of mouse, rabbit, and rat liver, lung, and kidney can metabolize compounds containing thiol, sulfide, thioamide, secondary and tertiary amine, hydrazine, and phosphine substituents. Therefore, this enzyme from these common experimental animals has catalytic capabilities similar to those of the well-characterized porcine liver enzyme. True allosteric activation by n-octylamine does not appear to be a property of either the mouse, rabbit, or rat liver enzymes, but is a property of the pig liver and mouse lung enzymes. The microsomal pulmonary flavin-containing monooxygenase of the rabbit has some unique substrate preferences which differ from the mouse lung enzyme. Both the rabbit and mouse pulmonary enzymes have recently been shown to be distinct enzyme forms. However, the rat pulmonary flavin-containing monooxygenase appears to be catalytically identical to the rat liver enzyme, and does not have any of the unusual catalytic properties of either the rabbit or mouse lung enzymes. Enzyme activity of mouse, rabbit, and rat kidney microsomes is qualitatively similar to the hepatic activities. Substrates which saturate the microsome-bound flavin-containing monooxygenase at 1.0 mM, including thiourea, thioacetamide, methimazole, cysteamine, and thiobenzamide, are metabolized at common maximal velocities. This suggests that the kinetic mechanism of the native enzyme is similar to that established for the isolated porcine liver enzyme in that the rate-limiting step of catalysis occurs after substrate binding, and that all substrates capable of saturating the microsomal enzyme should be metabolized at a common maximal velocity.     

Purification and characterization of cholyl-CoA: taurine N-acetyltransferase from the liver of domestic fowl (Gallus gallus).

The enzymological basis for the ability of mammalian liver to conjugate bile acids with both glycine and taurine, and for non-mammalian liver to make only taurine conjugates, was investigated. The taurine-conjugating enzyme has been purified 1200-fold from the liver of domestic fowl and its properties compared with those of the glycine/taurine-conjugating enzyme from bovine liver [Czuba & Vessey (1980) J. Biol. Chem. 255, 5296-5299]. The enzyme from both species followed a Ping Pong mechanism. The enzymes were also similar with respect to their affinity for taurine, although the enzyme from domestic fowl would not bind glycine. The affinity of both for cholyl-CoA was quite similar, too, and both enzymes were inhibited reversibly by p-mercuribenzoate. The enzymes, however, were quite different in size. The enzyme from domestic fowl had a mol.wt. of 63000-65000 by both gel filtration and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. This is approx. 15 000 mol.wt. units larger than the enzyme from bovine liver, and suggests a loss of genome over the course of evolution as the basis for the altered specificity at the amino-acid binding site.     

Development of phenylalanine hydroxylase activity in rat kidney

The developmental pattern of phenylalanine hydroxylase was studied in rat kidney and compared with that of liver from the same animal. Traces of activity were observed from 19 to 21 days of gestation in the liver and on Day 20 and 21 of gestation in the kidney. Significant amounts of activity were noticed on Day 22 of gestation. Kidney on Day 21 of gestation showed slightly higher values than that seen in corresponding liver. Fetal liver and kidney showed about 30% and 90% activity of newborn animals, respectively. Both synthetic cofactor and organic reductant were necessary for optimal activity of liver and kidney enzymes. pH studies showed an optimum at pH 7.0 in both liver and kidney. Storage at ?15 °C resulted in loss of activity in both liver and kidney to the same extent at a given time. No evidence of inhibition of either liver or kidney enzyme by phenylalanine was noticed up to a concentration of 4 μmoles per assay. Heat denaturation studies at 50 °C showed the kidney enzyme to be slightly less stable than the liver enzyme though a similar pattern was observed in both tissues at all ages studied.     

Physiological significance of catalase and glutathione peroxidases,and in vivo peroxidation,in selected tissues of the toadDiscoglossus pictus (Amphibia) during acclimation to normobaric hyperoxia

1. Various parameters related to oxidative stress were measured in adult Discoglossus pictus acclimated for 15 days to either normoxia or hyperoxia (PO2 = 710 mmHg). 2. Total weight of the toads and total and relative wet weight of liver, kidneys, lungs and heart were not changed by hyperoxic acclimation. 3. In vivo tissue peroxidation increased in lung, decreased in skeletal muscle, and was not changed in liver, kidney, heart and skin after hyperoxic exposure. 4. Hyperoxic acclimation increased catalase activities in the lung, liver, kidney and heart but not in skeletal muscle and skin. 5. Liver showed higher GSH-peroxidase activity with cumene-OOH than with H2O2 as substrate, whereas lung, skeletal muscle and skin presented similar GSH-peroxidase activities with both substrates. 6. GSH-peroxidase activities did not change between hyperoxic and normoxic animals in liver, lung, skeletal muscle and skin. 7. These results show that catalase, not GSH-peroxidase, is the principal H2O2 detoxifying enzyme involved in the adaptation of D. pictus to hyperoxia.     

Comparison of dipeptidyl peptidase IV prepared from pig liver and kidney

Dipeptidyl peptidase IV (dipeptidylpeptide hydrolase, EC 3.4.14.-) has been purified from the microsomal fraction of pig liver, using an immunoaffinity chromatography, and its properties compared with those of the enzyme purified from pig kidney. The amino acid compositions of both enzymes were similar. The same kinds of carbohydrates were found in both enzymes, but there were differences in the molar concentrations of individual sugars. The liver enzyme had greater concentrations of mannose, fucose and sialic acid than the kidney enzyme, while the concentrations of galactose and glucosamine were greater in the kidney enzyme. The carbohydrates accounted for approx. 18.3 and 22.7% of the weight of the kidney and liver enzymes, respectively. The pH optima, molecular weights, substrate specificities and Km values of the two enzymes and the effects of diisopropylfluorophosphate on their activities were nearly identical. The liver enzyme was heat- and pH-sensitive, but not attacked by proteinases.     

Expression of functional human glutaminase in baculovirus system: affinity purification, kinetic and molecular characterization

Glutaminase catalyzes the hydrolysis of glutamine yielding stoichiometric amounts of glutamate plus ammonium ions. In mammals, there are two different genes encoding for glutaminase, known as liver (L) and kidney (K) types. The human L-type isoform expressed in baculovirus yielded functional recombinant enzyme in Sf9 insect cells. A novel affinity chromatography method, based on its specific interaction with a PDZ protein, was developed for purification. Kinetic constants were determined for the purified human isozyme, which showed an allosteric behaviour for glutamine, with a Hill index of 2.7 and S(0.5) values of 32 and 64 mM for high and low P(i) concentrations, respectively. Whereas the protein showed a low P(i) dependence typical for L-type glutaminases, the enzyme was unexpectedly inhibited by glutamate, a kinetic characteristic exclusive of K-type isozymes, and was slightly activated by ammonia, unlike the classical liver enzymes which show an absolute dependence on ammonia. Subcellular fractionation demonstrates that recombinant human glutaminase was targeted to both mitochondria and nucleus, and in both locations the protein was catalytically active. This is the first report of the expression of a functional L-type mammalian glutaminase enzyme. The study also provides a simple and efficient method for affinity purification of the recombinant enzyme. Moreover, the data imply that this human enzyme may represent a new isoform different from classical kidney and liver isozymes.     

Molecular differences between rat-liver and rat-kidney biliverdin reductase. Implications for their in vivo regulation

Rat-liver biliverdin reductase exists in two molecular forms. The major form 1 has a molecular mass of 34 kDa, while the minor form 2 has a molecular mass of 56 kDa. Form 1 was converted into a second major form (form 3) with a molecular mass of 68 kDa by a NAD+-dependent peroxisomal dehydrogenase which was induced under conditions of oxidative stress [Frydman, R. B., Tomaro, M. L., Awruch, J. & Frydman, B. (1984) Biochem. Biophys. Res. Commun. 121, 249]. Molecular form 1 from rat kidney was not affected by the dehydrogenase, and a structural explanation for this difference was therefore sought. Both form 1 biliverdin reductases, isolated from rat liver and kidney, were purified to homogeneity using affinity chromatography, FPLC and HPLC techniques. The homogeneous enzymes were found to be identical when compared by their HPLC retention times, amino acid compositions and electrophoretic behaviour on polyacrylamide gels under non-denaturing conditions and on SDS/polyacrylamide gels. On HPLC analysis the peptides resulting from the CNBr cleavage were found to be the same for both enzymes, when either the native enzymes or their thioethylpyridine derivatives were compared. When the HPLC fingerprints of the tryptic digests were compared, they were found to be very similar, except for a peptide eluting at 31.60 min in the liver digest and at 23.60 min in the kidney digest. When the enzyme from both origins was alkylated with 4-dimethylaminoazobenzene-4'-iodoacetamide and then digested with trypsin, the HPLC fingerprints of the alkylated cysteine-carrying peptides were almost identical, except for a peptide with a retention time of 19.03 min in the liver digest and of 18.19 min in the kidney digest. The liver reductase was not amenable to Edman degradation suggesting a block at the NH2-terminus; in the kidney enzyme, however, it was free and an NH2-terminal sequence of 12 amino acids could be determined. The liver enzyme was found to be more sensitive toward p-hydroxymercuriphenyl sulfonate than the kidney enzyme.     

A rapid purification procedure for camel kidney glutathione S-transferase

Glutathione S-transferase was isolated from supernatant of camel kidney homogenate centrifugation at 37,000 xg by glutathione agarose affinity chromatography. The enzyme preparation has a specific activity of 44 mumol/min/mg protein and recovery was more than 85% of the enzyme activity in the crude extract. Glutathione agarose affinity chromatography resulted in a purification factor of about 49 and chromatofocusing resolved the purified enzyme into two major isoenzymes (pI 8.7 and 7.9) and two minor isoenzymes (pI 8.3 and 6.9). The homogeneity of the purified enzyme was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration on Sephadex G-100. The different isoenzymes were composed of a binary combination of two subunits with molecular weight of 29,000 D and 26,000 D to give a native molecular weight of 55,000 D. The substrate specificities of the major camel kidney glutathione S-transferase isoenzymes were determined towards a range of substrates. 1-chloro-2,4-dinitrobenzene was the preferred substrate for all the isoenzymes. Isoenzyme III (pI 7.9) had higher specific activity for ethacrynic acid and isoenzyme II (pI 8.3) was the only isoenzyme that exhibited peroxidase activity. Ouchterlony double-diffusion analysis with rabbit antiserum prepared against the camel kidney enzyme showed fusion of precipitation lines with the enzymes from camel brain, liver and lung and no cross reactivity was observed with enzymes from kidneys of sheep, cow, rat, rabbit and mouse. Different storage conditions have been found to affect the enzyme activity and the loss in activity was marked at room temperature and upon repeated freezing and thawing.     

A comparison of the binding of endothelin to various tissues from spontaneously hypertensive and Wistar-Kyoto rats

The binding affinities for endothelin-1 and endothelin-3 to membrane preparations of various tissues of spontaneously hypertensive rats and normotensive Wistar-Kyoto rats were compared by competition binding of the peptides with [¹²⁵I]endothelin-1. Endothelin-1 binding data obtained using membrane preparations from brain, heart, kidney, liver, lung and spleen of both strains were better fit with a one-site model. The brain tissue demonstrated the highest affinity for endothelin-1 in both strains with the same IC₅₀ of 0.11 nM, while the kidney and lung tissues showed the lowest affinities in both strains with IC₅₀ values ranged between 1.4 and 4.1 nM. Only the kidney tissues of these two strains showed a statistically significant difference in binding affinities for endothelin-1; the IC₅₀ values were 1.4 ± 0.1 nM (mean ± SE, N = 3) and 3.2 ± 0.4 nM (n = 4) for the spontaneously hypertensive and normotensive rats, respectively. Endothelin-3 binding data obtained using membrane preparations from brain, kidney and lung of both strains were also better fit with a one-site model. In contrast, a two-site model was more suitable for analyzing endothelin-3 binding results obtained using membrane preparations from heart, liver and spleen of both strains. Again, only the kidney tissues of the two strains showed a statistically significant difference in binding affinities for endothelin-3. The ratio of IC₅₀ value of the major endothelin-3 binding site to that of endothelin-1 in each tissue varied from approx. 1.5 in brain, kidney and liver to greater than 500 in heart and spleen of both strains. Scatchard analysis of saturation binding data showed that [¹²⁵I]endothelin-1 bound to a single class of binding sites in brain, heart, liver and spleen of both rat strains and in kidney of the spontaneously hypertensive rats. Specific binding to the kidney membrane preparation of the normotensive rats was not saturable at radioligand concentrations up to about 2 nM. These results suggest that the tissues of both strains investigated have different affinities as well as different selectivities for endothelin-1 and endothelin-3. Furthermore, kidney is the only tissue examined which showed higher binding affinity in the spontaneously hypertensive rats than that of the normotensive ones.     

A Rapid Purification Procedure for Camel Kidney Glutathione S-Transferase

Glutathione S-transferase was isolated from supernatant of camel kidney homogenate centrifugation at 37, 000 xg by glutathione agarose affinity chromatography. The enzyme preparation has a specific activity of 44 μ;mol/min/mg protein and recovery was more than 85% of the enzyme activity in the crude extract. Glutathione agarose affinity chromatography resulted in a purification factor of about 49 and chromatofocusing resolved the purified enzyme into two major isoenzymes (pI 8.7 and 7.9) and two minor isoenzymes (pI 8.3 and 6.9). The homogeneity of the purified enzyme was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration on Sephadex G-100.

The different isoenzymes were composed of a binary combination of two subunits with molecular weight of 29, 000 D and 26, 000 D to give a native molecular weight of 55, 000 D.

The substrate specificities of the major camel kidney glutathione S-transferase isoenzymes were determined towards a range of substrates. l-chloro-2, 4-dinltrobenzene was the preferred substrate for all the isoenzymes. Isoenzyme III (pI 7.9) had higher specific activity for ethacrynic acid and isoenzyme II (pI 8.3) was the only isoenzyme that exhibited peroxidase activity. Ouchterlony double-diffusion analysis with rabbit antiserum prepared against the camel kidney enzyme showed fusion of precipitation lines with the enzymes from camel brain, liver and lung and no cross reactivity was observed with enzymes from kidneys of sheep, cow, rat, rabbit and mouse.

Different storage conditions have been found to affect the enzyme activity and the loss in activity was marked at room temperature and upon repeated freezing and thawing.     

Characterization of ferrochelatase in kidney and erythroleukemia cells

Ferrochelatase from bovine kidney mitochondria has been purified 1600-fold with a 6.5% yield, exhibiting a specific activity of 490 nmol mesoheme formed/mg of protein per min. The Km values for mesoporphyrin IX and protoporphyrin IX with iron were 12.5 and 12.7 microM, respectively. The Km values for iron and zinc with mesoporphyrin IX were 3.51 and 3.17 microM, respectively. The purified enzyme showed a single band with an apparent molecular mass of 42,000 daltons (42 kDa) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The rabbit antibody against the purified enzyme markedly inhibited activities of the enzyme from both the kidney and liver. Immunoblot analysis showed that the antibody reacted with the renal as well as the hepatic enzymes showing the same molecular weight. Peptide mapping with trypsin or alpha-chymotrypsin showed that digested peptides of renal enzyme were similar to those of hepatic enzyme. Ferrochelatase activity in mouse erythroleukemia (MEL) cells increased in parallel with an increase of heme synthesis by treatment with dimethylsulfoxide. Using immunoblotting techniques, the amount of the enzyme in the MEL cells has been shown to increase by the induction, showing a molecular mass of 41 kDa which was the same as that of the mouse hepatic enzyme. Comparative structural analysis of the enzyme of MEL cells and that of mouse liver by peptide mapping showed that the partial digestive peptides of both enzymes exhibited a similar pattern. These results strongly suggest that ferrochelatase in kidney, liver and erythroid cells can be of one type.     

Mouse liver cytoplasmic malate dehydrogenase: rapid isolation, characterization and immunochemical comparison with hepatoma enzyme.

Cytoplasmic NAD-dependent malate dehydrogenase is decreased in activity in three transplantable mouse hepatomas compared to the activity of this enzyme in liver tissue. This enzyme is composed of several molecular forms of similar size which differ slightly in charge; the total activity and the discernible number of forms of the enzyme are decreased in both hepatoma and fetal liver. Mixing experiments suggest the absence of a significant quantity of unbound inhibitor of enzyme activity in the tumor or an activator in the liver. The liver cytoplasmic enzyme was purified to homogeneity by a relatively rapid method using Blue Sepharose affinity chromatography, which results in a good yield and high specific activity of the enzyme. Cytoplasmic and mitochondrial enzymes bind and elute differentially from this affinity resin. Molecular weight, kinetic constants and amino acid composition of the cytoplasmic enzyme were determined. Monospecific antiserum to the cytoplasmic enzyme has been produced in a goat and used to demonstrate a lack of immunological cross-reactivity between the mitochondrial and cytoplasmic enzyme. The tumor and liver cytoplasmic enzymes possess similar, if not identical, immunological determinants. Immunotitration experiments have been used to demonstrate that liver and hepatoma enzyme are identical in specific activity. Thus, the reduction in level of cytoplasmic enzyme in hepatoma is due to a decrease in the number of molecules per tissue mass.     

Regulation of aspartate aminotransferase activity associated with change of pyridoxal phosphate level.

The activities of aspartate aminotransferase (EC 2.6.1.1) in the cytosol fractions of the liver and kidney of rats fed pyridoxine-deficient or control diet for 3 weeks were determined. In the absence of pyridoxal phosphate, the activities in the liver and kidney preparations of deficient rats were both abnormally low. The activity in the kidney fraction of deficient rats was restored to almost the control level by addition of pyridoxal phosphate, whereas that of the liver was only partially restored. The antigen activity, however, measured using anti-aspartate aminotransferase, was similar in liver fractions from deficient and control rats. These findings suggest the existence of a form of transaminase with little or no activity in the liver of deficient rats. The properties of the crude enzymes from deficient and control rats were indistinguishable by immunodiffusion, and the enzymes had the same subunit size and heat stability under the conditions tested. However, purified enzyme from deficient rat liver had a different specific activity and absorption spectrum from purified enzyme from normal liver.     

The flavin-containing monooxygenase in mouse lung: Evidence for expression of multiple forms

The flavin-containing monooxygenase (FMO) was purified from mouse lung microsomes. On SDS-PAGE, the purified enzyme separated as two bands, a major band of 58,000 daltons and a minor band of 59,000 daltons. Antibodies to mouse liver FMO cross-reacted with both bands in the purified preparations, whereas antibodies to rabbit lung FMO cross-reacted only with the major band. In microsomal preparations the major band was recognized by both antibodies, but neither antibody detected the minor band in microsomes. A cDNA encoding the pig liver FMO hybridized with mRNA isolated from mouse liver, kidney, and lung, whereas cDNA encoding the rabbit lung FMO hybridized only with mouse lung and kidney mRNA. Thermal stability studies showed that the FMO preparation purified from mouse lung consisted of a heat-stable and a heat-labile component. The heat-labile component of lung FMO was inhibited competitively by imipramine, whereas the heat-stable component was insensitive to the presence of imipramine. Immunoprecipitation of purified mouse lung FMO with anti-rabbit lung FMO completely removed the protein band reactive to anti-rabbit lung FMO while leaving reactivity to anti-liver FMO. The catalytic and immunochemical differences seen between FMO from rabbit lung and mouse lung appear to result from the expression of at least two forms of FMO in the mouse lung, one similar to the rabbit pulmonary form and one similar to the major mouse liver form of FMO.     

Purification and properties of cytosolic alanine aminotransferase from the liver of two freshwater fish, Clarias batrachus and Labeo rohita

Cytosolic alanine aminotransferase (c-AAT) was purified up to 203- and 120-fold, from the liver of two freshwater teleosts Clarias batrachus (air-breathing, carnivorous) and Labeo rohita (water-breathing, herbivorous), respectively. The enzyme from both fish showed similar elution profiles on a DEAE-Sephacel ion exchange column. SDS-PAGE of purified enzymes revealed two subunits of 54 and 56 kDa, in both fish. The apparent Km values for l-alanine were 18.5+/-0.48 and 23.55+/-0.60 mM, whereas for 2-oxoglutarate the Km values were observed to be 0.29+/-0.023 and 0.33+/-0.028 mM for the enzyme from C. batrachus and L. rohita, respectively. With l-alanine as substrate, aminooxyacetic acid was found to act as a competitive inhibitor with KI values of 6.4 x 10(-4) and 3.4 x 10(-4) mM with c-AAT of C. batrachus and L. rohita, respectively. However, when 2-oxoglutarate was used as substrate, aminooxyacetic acid showed uncompetitive inhibition with similar KI values for purified c-AAT from both fish. Temperature and pH profiles of the enzyme did not show any marked differences between the two fish examined. These results suggest that liver c-AAT, isolated from these two fish species adapted to different modes of life, remain unaltered structurally. However, at the kinetic level, liver c-AAT from C. batrachus exhibits significantly higher affinity for the substrate l-alanine and decreased affinity for its metabolic inhibitor, in comparison to that of the enzyme purified from L. rohita. Such functional changes seem to be of physiological significance and also provide preliminary evidence for subtle changes in the enzyme as a mark of metabolic adaptation in the fish to different physiological demands.     

Metabolic activation of food mutagens by liver and lung enzymes from rats, pigs and oxen

Mutagenic activation of the 3 cooked food mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) was compared in liver and lung enzyme preparations from oxen, pigs and rats. Liver preparations from oxen were the most efficient in activating the mutagens, while the rat enzymes were more active than those from pigs. The different cooking mutagens showed different mutagenic potential. MeIQ was the most potent mutagen, followed by IQ and MeIQx in descending order. In oxen, MeIQx was as potent as IQ. The activation with the lung enzymes was 2-3 orders of magnitude lower than with liver. Furthermore, species differences in mutagenic activation with lung enzymes were small compared with liver enzymes. In lung preparations the differences between IQ and MeIQ were small, but in all 3 animal species the mutagenicity of MeIQx was 1 order of magnitude lower than that of the other 2 mutagens.     

Some properties of phosphofructokinase from kidney cortex and their relation to glucose metabolism

1. Phosphofructokinase from rat kidney cortex has been partially purified by using a combination of isoelectric and ammonium sulphate precipitation. This preparation was free of enzymes which interfered with the measurement of either product of phosphofructokinase. 2. At concentrations greater than the optimum, ATP caused inhibition which was decreased by raising the fructose 6-phosphate concentration. This suggested that ATP reduced the affinity of phosphofructokinase for the other substrate. Citrate potentiated the ATP inhibition. 3. AMP and fructose 1,6-diphosphate relieved the inhibition by ATP or citrate by increasing the affinity of the enzyme for fructose 6-phosphate. 4. K(+) is shown to stimulate and Ca(2+) to inhibit phosphofructokinase. 5. The similarity between the complex properties of phosphofructokinase from kidney cortex and other tissues (e.g. cardiac and skeletal muscle, brain and liver) suggests that the enzyme in kidney cortex tissue is normally subject to metabolic control, similar to that in other tissues.     

Distribution of two urinary ribonuclease-like enzymes in human organs and body fluids

In order to determine the distribution of two human urinary RNase (RNase Us and RNase UL)-like enzymes in human tissues and body fluids, enzyme immunoassay systems were established using rabbit anti-RNase sera. The sensitivity of the assay systems was of similar order to that of radioimmunoassay systems previously reported. In the enzyme immunoassay, the cross reactivities of anti-RNase UL serum towards RNase Us, bovine kidney RNase K2, bovine RNase A, and bovine seminal RNase Vs were less than 1%. The cross reactivity of anti-RNase Us-serum towards RNase UL was less than 0.5% and cross reactivities were minimal for RNase A, RNase K2, and RNase Vs. The RNase levels in human organs and body fluids were measured by enzyme immunoassay. In milk, semen and saliva, only RNase UL-like enzyme was found. Both RNase Us- and RNase UL-like enzymes were found in kidney, stomach, and pancreas and the RNase Us/RNase UL ratios were 0.49, 1.35, and 0.34, respectively. In lung, liver, spleen, and leukocytes, most of the RNase activity was accounted for by RNase Us-like enzyme. The activity of RNase Us-like enzyme was especially high in lung, spleen, and leukocytes. The crude extracts of several tissues and body fluids were separated by phosphocellulose column chromatography and the contents of the two urinary RNase-like enzymes were determined by enzyme immunoassay. In stomach, kidney, pancreas, and serum, both enzymes were present in multiple forms. In spleen and lung, both the major RNase (RNase Us) and minor RNase (RNase UL) existed in two forms.(ABSTRACT TRUNCATED AT 250 WORDS)     

大白鼠DNA拓扑异构酶Ⅰ生物学性质的研究

对大白鼠组织作DNA拓扑弄构酶Ⅰ(拓扑酶Ⅰ)活力测定,见酶活力出现在胚胎早期,在胚胎发育过程及出生后不同年龄期,酶活力基本稳定;几种成年大鼠组织的酶活力彼此无显著差异;肝细胞再生及癌变,酶活力亦无显著变化。