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1.
Spermatozoa released from the seminiferous tubules are terminally differentiated cells with no known synthetic activity. Their components are synthesized in the spermatogenic cells during spermatogenesis. In this study, we report the characterization and immunolocalization of beta-glucuronidase in mouse testicular germ cells and spermatozoa. The enzyme is an exoglycohydrolase with dual localization, being present in lysosomes and endoplasmic reticulum of several mouse and rat tissues. The purified germ cell preparations (spermatocytes, round spermatids, and condensed/elongated spermatids) when assayed for beta-glucuronidase activity showed that the spermatocytes contained five times more enzyme activity per cell than the spermatids. Polyacrylamide gel electrophoresis, carried out under native and denaturing conditions, demonstrated that the germ cells express only the lysosomal form of the enzyme (pI 5.5-6.0) with a subunit molecular mass of 74 kDa. Immunocytochemical studies revealed a positive reaction in the Golgi membranes, Golgi-associated vesicles, and lysosomes of late spermatocytes (pachytene spermatocytes) and a stage-specific localization during spermiogenesis. The forming or formed acrosome of the elongated spermatids (stages 9-16) and epididymal spermatozoa was highly immunopositive. Comparison of immunoprecipitation curves and kinetic properties of the enzyme present in spermatocytes and spermatozoa revealed no major differences. Taken together, our results demonstrate that beta-glucuronidase activities present in the lysosomes of spermatocytes and the sperm acrosome are kinetically and immunologically similar. 相似文献
2.
Three differently modified forms of beta-glucuronidase are known to exist: a microsomal enzyme form (M) existing in tissues where egasyn, a second microsomal protein, is present; and an acidic (La; complex-type oligosaccharide) and a basic (Lb; non-complex type oligosaccharide) lysosomal form which occur in all mouse tissues. Lb predominates in tissues containing microsomal beta-glucuronidase, La in those lacking it. In pulse-labelling experiments using mouse strain C57BL/6 liver containing egasyn (Eg+/Eg+) and microsomal enzyme, about half of the newly synthesized beta-glucuronidase was processed to the microsomal enzyme form, which was evidently further processed to Lb, and about half directly to La. In contrast, in liver of the congenic line C57BL/6.YBR Es-1b Eg0 that lacks egasyn (Eg0/Eg0) and microsomal enzyme, most of the labelled beta-glucuronidase was processed to La, and only a minor portion to Lb. Newly synthesized enzyme appeared first in microsomal, then in light and heavy lysosomal fractions of Eg+/Eg+ liver. In Eg0/Eg0 liver, no labelled enzyme was measurable in the microsomes, but it appeared rapidly in both types of lysosomes. Taken together these findings indicate that the microsomal enzyme form serves as a precursor of Lb, and that La is synthesized independently. The apparent half-life of La is only two-thirds that of Lb; this fact accounts for the reduced beta-glucuronidase activity in Eg0/Eg0 liver, which contains La as the predominant form. 相似文献
3.
C A Brown 《The journal of histochemistry and cytochemistry》1978,26(1):22-27
Colon tumors induced with azoxymethane in male Fischer rats were cytochemically analyzed for beta-glucuronidase using naphthol AS-B1 glucuronide (6-bromo-2-hydroxy-3-naphthoyl-O-anisidine) as a substrate and hexazonium pararosanin as a diazo reagent. This method effectively localizes the bulk of beta-glucuronidase in the surface epithelium, the lamina propria and in the endothelial cells of the lymphoid sinuses and postcapillary venules. Polypoid lesions, adenocarcinomas and mucinous adenocarcinomas show no difference in the amount or in the localization of beta-glucuronidase; however, mucinous adenocarcinomas show a slight increase in the amount of beta-glucuronidase. The few tumors that did metastasize to lymph nodes did not show any difference in their enzyme patterns. Intestinal crypts that show a change in size and shape have a definite increase in beta-glucuronidase activity. An increase in the activity of this enzyme can also be seen in well defined neoplasms as opposed to normal areas of the colon. 相似文献
4.
—The properties and subcellular localization of type I (nitrophenyl) and type II (nitrocatechol) arylsulphatases were investigated in brain tissue of the rat, and optimal assay conditions were established. Sulphate, phosphate and sulphite ions inhibited the nitrocatechol sulphatases; nitrophenyl sulphatase was inhibited only by sulphite. The presence of latent enzyme activity was demonstrated for the nitrocatechol sulphatases, beta-glucuronidase, and beta-glycerophosphatase in rat and mouse brain homogenates. These hydrolases were highly sensitive to mechanical and osmotic damage; and Triton X-100 was very effective in releasing their latent (bound) activities, a finding suggestive of a lysosomal localization. Activity of nitrophenyl sulphatase was unaffected by osmotic changes or Triton X-100, characteristics suggesting a membranous association for this enzyme. Total activity of nitrophenyl sulphatase was approximately twice as great in canine gray matter as in canine white matter; the converse obtained for beta-glucuronidase activity. Values for total enzymic activity of the nitrocatechol sulphatases in canine white and gray matter were similar. Fractionation of homogenates from rat brain by differential centrifugations and separation of crude mitochondrial fractions by sucrose density gradient centrifugations revealed the following: (1) most of the nitrocatechol sulphatase activity (93 per cent) and all of the nitrophenyl sulphatase activity were sedimentable; (2) crude mitochondrial fractions exhibited the highest relative specific activity (RSA = 1·38) for the nitrocatechol sulphatases, whereas microsomal fractions displayed the highest RSA for nitrophenyl sulphatase (1·89); (3) the lightest fraction (A + B) and the densest fraction (E) from the sucrose density gradient contained most of the activity for both the type I and type II arylsulphatases, whereas the RSA of cytochrome oxidase was greatest in the intermediate density regions (fractions C and D); (4) the highest RSA for beta-glucuronidase and beta-glycerophosphatase occurred in gradient fraction C; (5) appreciable activity of beta-glycerophosphatase was found in a nerve ending fraction (M3). It is suggested that the hydrolases in heterogeneous tissue like brain might be associated with lysosomal particles of differing enzyme compositions and varying populations, and that the data on distribution lend credence to the concept of bimodal and possible trimodal particle affinity for the hydrolases of brain tissues. 相似文献
5.
Failure of Urinary Beta-Glucuronidase Activity to Localize the Site of Urinary Tract Infection 总被引:1,自引:0,他引:1
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Allan R. Ronald Frederic Silverblatt Hugh Clark Ralph E. Cutler Marvin Turck 《Applied microbiology》1971,21(6):990-992
The differentiation of renal from bladder bacteriuria is difficult on clinical grounds alone. To evaluate the correlation between site of infection and urinary beta-glucuronidase activity, 46 patients with well documented recurrent bacteriuria were studied by bilateral ureteral catheterization. Urinary beta-glucuronidase activity was also determined in 46 control subjects. In general, asymptomatic patients with renal bacteriuria, either unilateral or bilateral, had levels of enzyme activity in their urine comparable to patients with infection confined to the bladder and to normals. Only 4 of 25 patients with renal bacteriuria had significant elevations of urinary beta-glucuronidase. After localization of infection, 9 of 10 patients treated with kanamycin, a potentially nephrotoxic drug, developed significant elevations of urinary beta-glucuronidase. The results of these studies indicate that determination of beta-glucuronidase activity in urine is not useful in predicting the site of infection in patients with bacteriuria but may find a role in screening for early nephrotoxicity. 相似文献
6.
GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. 总被引:63,自引:1,他引:62
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We have used the Escherichia coli beta-glucuronidase gene (GUS) as a gene fusion marker for analysis of gene expression in transformed plants. Higher plants tested lack intrinsic beta-glucuronidase activity, thus enhancing the sensitivity with which measurements can be made. We have constructed gene fusions using the cauliflower mosaic virus (CaMV) 35S promoter or the promoter from a gene encoding the small subunit of ribulose bisphosphate carboxylase (rbcS) to direct the expression of beta-glucuronidase in transformed plants. Expression of GUS can be measured accurately using fluorometric assays of very small amounts of transformed plant tissue. Plants expressing GUS are normal, healthy and fertile. GUS is very stable, and tissue extracts continue to show high levels of GUS activity after prolonged storage. Histochemical analysis has been used to demonstrate the localization of gene activity in cells and tissues of transformed plants. 相似文献
7.
Quercetin glucuronides are the main circulating metabolites of quercetin in humans. We hypothesise that the potential availability of the aglycone within tissues depends on the substrate specificity of the deconjugating enzyme beta-glucuronidase towards circulating flavonoid glucuronides. Human tissues (small intestine, liver and neutrophils) exhibited beta-glucuronidase against quercetin glucuronides. The various quercetin glucuronides were deconjugated at similar rates, but liver cell-free extracts were the most efficient and the activity was completely inhibited by saccharo-1,4-lactone (a beta-glucuronidase inhibitor). Furthermore, pure recombinant human beta-glucuronidase hydrolysed various flavonoid glucuronides, with a 20-fold variation in catalytic efficiency (k(cat)/K(m)=1.3x10(3) M(-1) s(-1) for equol-7-O-glucuronide and 26x10(3) M(-1) s(-1) for kaempferol-3-O-glucuronide). Similar catalytic efficiencies were obtained for quercetin O-glucuronides substituted at different positions. These results show that flavonoid glucuronides can be deconjugated by microsomal beta-glucuronidase from various human cells. 相似文献
8.
Expression of active human beta-glucuronidase in Sf9 cells infected with recombinant baculovirus 总被引:2,自引:0,他引:2
Antibody directed enzyme prodrug therapy (ADEPT) using glucuronide prodrugs is an experimental approach to reduce systemic toxicity of anti-cancer agents. Bioactivation of such prodrugs is achieved by fusion proteins consisting of targeting moieties (e.g. ligands of tumor specific antigens) and human beta-glucuronidase. In order to test a large panel of possible beta-glucuronidase fusion proteins for their applicability in ADEPT, an easy, rapid and high-yield expression system like the baculovirus/insect cell expression system would be needed. A prerequisite for using such fusion proteins is functional and biochemical characterization of human beta-glucuronidase expressed in baculovirus-infected insect cells. Therefore, recombinant human beta-glucuronidase was expressed in Sf9 insect cells and characterized at the protein and functional level. As shown by Western blot analysis the recombinant enzyme consists of dimers with their monomers being linked via disulfide bonds. Posttranslational modifications of the monomers seem to be different as compared with mammalian cells or tissues. The enzyme is functionally active in cleaving the substrates 5-bromo-4-chloro-3-indolyl-beta-D-glucuronic acid, 4-methylumbelliferyl-beta-D-glucuronide and the glucuronide prodrug HMR 1826, respectively, with similar enzyme kinetic parameters as those found in human tissues. Our data demonstrate that beta-glucuronidase expressed in Sf9 cells displays the same enzymatic features as the protein expressed in mammalian cells. Therefore, we suggest that beta-glucuronidase fusion proteins produced in this cell line will be valuable tools for testing a large panel of various targeting moieties in human tumor xenograft models or may be used for ADEPT in man. 相似文献
9.
Purification and immunological quantification of rat liver lysosomal glycosidases. 总被引:1,自引:0,他引:1
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Although lysosomal enzyme activities are known to vary in response to numerous physiological and pharmacological stimuli, the relationship between lysosomal enzyme activity and enzyme concentration has not been systematically studied. Therefore we developed radioimmunoassays for two lysosomal glycosidases in order to determine lysosomal enzyme concentration. beta-Galactosidase and beta-glucuronidase were purified from rat liver 2780-fold and 1280-fold respectively, by using differential centrifugation, affinity chromatography, ion-exchange chromatography and molecular-sieve chromatography. Polyclonal antibodies to these enzymes were raised in rabbits, and two radioimmunoassays were established. Antibody specificity was shown by: (i) selective immunoprecipitation of enzyme activity; (ii) identical bands of purified enzyme on SDS/polyacrylamide-gel electrophoresis and immunoelectrophoresis; (iii) single immunoreactive peaks in molecular-sieve chromatography experiments. Sensitivities of the assays were such that 15 ng of beta-galactosidase and 45 ng of beta-glucuronidase decreased the ratio of bound to free radiolabel by 50%; minimal detectable amounts of immunoreactive enzymes were 2 ng and 10 ng respectively. The assays were initially used to assess the effects of physiological perturbations (i.e. fasting and age) on enzyme concentrations in rat liver; these experiments showed that changes in enzyme concentrations do not always correlate with changes in enzyme activities. This represents the first report of radioimmunoassays for lysosomal glycosidases. The results suggest that these radioimmunoassays provide useful technology for the study of regulatory control mechanisms of the concentrations of lysosomal glycosidases in mammalian tissues. 相似文献
10.
Genetic variation for enzyme structure and systemic regulation in two new haplotypes of the beta-glucuronidase gene of Mus musculus castaneus 总被引:5,自引:0,他引:5
Two new haplotypes of the [Gus] gene complex have been characterized following their transfer from Mus musculus castaneus, where they were found, to a C57BL/6J genetic background. The [GUS]CS haplotype carries a new structural allele, Gus-scs, coding for enzyme with decreased thermolability and lacking an antigenic site present in other beta-glucuronidase allozymes. The [Gus]CL haplotype carries another new structural allele, Gus-scl, that codes for enzyme with increased thermolability and possessing the antigenic site. Both CS and CL beta-glucuronidase have the same catalytic activity/molecule as the standard B allozyme from C57BL/6J mice. Mice carrying either the [Gus]CS or [Gus]CL haplotype have reduced enzyme activity in all tissues examined at all stages of development. The reduced enzyme activity is partially accounted for by reduced rates of enzyme synthesis, and the remainder probably results from increased rates of enzyme turnover. beta-Glucuronidase mRNA levels in these mice were not reduced suggesting that the observed reduction in enzyme synthesis is due to a decreased efficiency of translation for CS and CL mRNA. 相似文献
11.
Guard cell- and phloem idioblast-specific expression of thioglucoside glucohydrolase 1 (myrosinase) in Arabidopsis 总被引:9,自引:0,他引:9
Thioglucoside glucohydrolase 1 (TGG1) is one of two known functional myrosinase enzymes in Arabidopsis. The enzyme catalyzes the hydrolysis of glucosinolates into compounds that are toxic to various microbes and herbivores. Transgenic Arabidopsis plants carrying beta-glucuronidase and green fluorescent protein reporter genes fused to 0.5 or 2.5 kb of the TGG1 promoter region were used to study spatial promoter activity. Promoter activity was found to be highly specific and restricted to guard cells and distinct cells of the phloem. No promoter activity was detected in the root or seed. All guard cells show promoter activity. Positive phloem cells are distributed in a discontinuous pattern and occur more frequent in young tissues. Immunocytochemical localization of myrosinase in transverse and longitudinal sections of embedded material show that the TGG1 promoter activity reflects the position of the myrosinase enzyme. In the flower stalk, the myrosinase-containing phloem cells are located between phloem sieve elements and glucosinolate-rich S cells. Our results suggest a cellular separation of myrosinase enzyme and glucosinolate substrate, and that myrosinase is contained in distinct cells. We discuss the potential advantages of locating defense and communication systems to only a few specific cell types. 相似文献
12.
Kinetics of beta-glucuronidase induction by androgen. Genetic variation in the first order rate constant 总被引:3,自引:0,他引:3
Measurements of enzyme activity, rates of protein synthesis, and mRNA activity suggest that the induction of beta-glucuronidase in mouse kidney in response to androgen is regulated at a pretranslational level. Following an initial lag period, the rate and extent of induction follow the rules of simple turnover kinetics and can be described in terms of a zero order rate constant for acquisition of mRNA activity (ka) and a first order rate constant for loss of activity (kb). Genetic variation in kb, described here for the first time, alters the half-time and extent of induction. Variation in kb is independent of previously described variation in ka and, unlike changes in ka, is not associated with change in the lag time. The DNA sequences determining kb, like those determining ka, are genetically linked to the structural gene for beta-glucuronidase. Following the removal of androgen, beta-glucuronidase activity, rate of synthesis, and mRNA activity all decline rapidly with half-lives of 1-2 days. Even in the most rapidly inducing strains, this is significantly faster than the half-time for induction determined by kb. Furthermore, genetic variation in kb does not affect the rate of de-induction. These facts suggest that kb may not describe the turnover of beta-glucuronidase mRNA, but rather the turnover of another step in the induction process. 相似文献
13.
Analysis of the expression of CLA1, a gene that encodes the 1-deoxyxylulose 5-phosphate synthase of the 2-C-methyl-D-erythritol-4-phosphate pathway in Arabidopsis
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Estévez JM Cantero A Romero C Kawaide H Jiménez LF Kuzuyama T Seto H Kamiya Y León P 《Plant physiology》2000,124(1):95-104
The discovery of the 2-C-methyl-D-erythritol-4-phosphate pathway for the biosynthesis of isoprenoids raises the important question of the nature and regulation of the enzymes involved in this pathway. CLA1, a gene previously isolated from Arabidopsis, encodes the first enzyme of the 2-C-methyl-D-erythritol-4-phosphate pathway, 1-deoxy-D-xylulose-5-phosphate synthase. We demonstrate this enzyme activity by complementation of the cla1-1 mutant phenotype and by direct enzymatic assays. Based on mRNA and protein expression patterns this enzyme is expressed mainly in developing photosynthetic and non-photosynthetic tissues. The beta-glucuronidase expression pattern driven from the CLA1 gene regulatory region supports the northern and protein data while also showing that this gene has some level of expression in most tissues of the plant. A mutation in the CLA1 gene interferes with the normal development of chloroplasts and etioplasts, but does not seem to affect amyloplast structure. Microscopic analysis also shows a pleiotropic effect of the CLA1 gene mutation in mesophyll tissue formation. 相似文献
14.
The intracellular localization of exogenously supplied human platelet beta-glucuronidase in cultured skin fibroblasts derived from a beta-glucuronidase-deficient patient was studied. Four cellular fractions were obtained by differential speed centrifugation. Following two days of incubation, the exogenously supplied enzyme exhibited a distribution pattern identical to that of endogenous beta-hexosaminidase. Disruption of membranes by freezing and thawing caused a 35% increase of the enzyme activity, thus indicating a latent activity following the internalization. This indicated localization in the lysosomal fractions. Longer incubation periods led to an intracellular shift of the engulfed enzyme from the lighter lysosomal fraction to heavier particles. Once located in the heavier fraction, the enzyme was relatively stable, and participated in the catabolism of 35S-labeled mucopolysaccharides which had accumulated in the lysosomes of these fibroblasts. A marked reduction in the accumulated mucopolysaccharides of the lysosomal fraction was observed following addition of the enzyme. This was accompanied by the formation of smaller sized molecules. 相似文献
15.
Summary Bone alkaline (AlP) and acid phosphatase (AcP) activities were simultaneusly demonstrated in tissue sections obtained from mice, rats, and humans. The method involved tissue fixation in ethanol, embedding in glycol methacrylate (GMA), and demonstration of AlP and AcP activities employing a simultaneous coupling azo dye technique using substituted naphthol phosphate as a substrate. AlP activity was demonstrated first followed by AcP activity. Both enzyme activities were demonstrated in tissue sections from bones fixed and/or stored in acetone or 70% ethanol for up to 14 days or stored in GMA for 2 months. AlP activity in tissue sections from bones fixed in 10% formalin, 2% glutaraldehyde, or formal-calcium, however, was markedly inhibited after 3–7 days and was no longer detectable after 14 days of fixation. Moreover, AlP activity was diminished in tissue sections from bones fixed in 70% ethanol or 10% formalin and subsequently demineralized in 10% EDTA (pH7) for 2 days, and the activity was completely abolished in tissue sections from bones subsequently demineralized in 5% formic acid: 20% sodium citrate (1:1, pH 4.2) for 2 days. Methyl methacrylate (MMA) embedding at concentrations above 66% completely inhibited AlP activity. AcP activity, however, was only partially inhibited by formalin, glutaraldehyde, or formal-calcium after 7 or 14 days of fixation or by MMA embedding and was unaffected by the demineralizing agent formic acid-citrate for 2 days. While AcP activity was preserved in bones fixed in formalin and subsequently demineralized in EDTA, the activity was completely abolished when EDTA demineralization was carried out on bones previously fixed in 70% ethanol. These results indicate that bone AlP and AcP activities can be demonstrated simultaneously in the same section using a simple tissue preparation technique and that the activities are retained in tissues fixed and/or stored in acetone, 70% ethanol or GMA, but are differentially inactivated by other fixatives studied, and by EDTA, formic acid-citrate, and MMA embedding.Abbreviations AcP
acid phosphatase
- AlP
alkaline phosphatase
- GMA
glycol methacrylate
- MMA
methyl methacrylate
- EDTA
ethylenediaminetetraacetic acid 相似文献
16.
Interaction of tumor and surrounding tissue of mice inoculated B16 melanoma variants in terms of enzyme activity 总被引:1,自引:0,他引:1
1. The interactions of B16-F1 and B16-F10 tumors with their surrounding tissues in terms of enzyme activities such as cathepsin B, hemoglobin(Hb)-hydrolase, acid phosphatase, beta-glucuronidase and plasminogen activator were investigated when said tumors proliferated locally and at secondary sites throughout the host's circulatory system. 2. In the case of B16-F1 and B16-F10 tumor cells proliferating under the skin, statistical differences were not detected between the enzyme activities of the skin surrounding the tumors and control skin, nor between B16-F1 and B16-F10 tumors, except for beta-glucuronidase. 3. In the case of B16-F1 and B16-F10 tumor cells metastasizing to lung, statistical differences were detected between numerous enzyme activities of the lung tissues surrounding the tumors and control lung tissue, and also between B16-F1 and B16-F10 tumors. 4. The activities of cathepsin B and acid phosphatase of lung tissue surrounding B16-F1 tumor were lower than those of the control lung. 5. beta-Glucuronidase activity of lung tissue surrounding B16-F10 tumor was higher than that of the control lung. 6. The activities of cathepsin B, Hb-hydrolase and beta-glucuronidase of the B16-F10 tumor were higher than those of the B16-F1 tumor. 7. Results indicate that metastasized B16 melanoma tumor cells interact with surrounding lung tissues, and that cathepsin B, Hb-hydrolase and beta-glucuronidase might play important roles in the metastasis of the malignant tumor. 相似文献
17.
Rat liver beta-glucuronidase. cDNA cloning, sequence comparisons and expression of a chimeric protein in COS cells. 总被引:3,自引:1,他引:2
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P P Powell J W Kyle R D Miller J Pantano J H Grubb W S Sly 《The Biochemical journal》1988,250(2):547-555
A cDNA for rat liver beta-glucuronidase was isolated, its sequence determined and its expression after transfection into COS cells studied. The deduced amino acid sequence of the rat liver clone showed 77% homology with that from the cDNA for human placental beta-glucuronidase and 47% homology with that deduced from the cDNA for Escherichia coli beta-glucuronidase. Several differences were found between the cDNA from rat liver and that previously reported from rat preputial gland. Only one change leads to an amino acid difference in the mature enzyme. A chimeric clone was constructed by using a fragment encoding the first 18 amino acid residues of the signal sequence from the human placental cDNA clone and a fragment from the rat clone encoding four amino acid residues of the signal sequence, all 626 amino acid residues of the mature rat enzyme, and all of the 3' untranslated region. After transfection into COS cells the chimeric clone expressed beta-glucuronidase activity that was specifically immunoprecipitated by antibody to rat beta-glucuronidase. The Mr value of 76,000 of the expressed gene product was characteristic of the glycosylated rat enzyme. It was proteolytically processed in COS cells to Mr 75,000 6 h after metabolic labelling. At least 50% of the expressed enzyme was secreted at 60 h post-transfection, but the secreted enzyme did not undergo proteolytic processing. These results provide evidence that the partial cDNA isolated from a rat liver library contains the complete coding sequence for the mature rat liver enzyme and that the chimeric signal sequence allows normal biosynthesis and processing of the transfected rat liver enzyme in COS cells. 相似文献
18.
J F de Sousa Júnior H B Nader C P Dietrich 《The Journal of biological chemistry》1990,265(33):20150-20155
A sulfatase acting upon chondroitin sulfate polymers, free of beta-glucuronidase and beta-N-acetylhexosaminidases, was isolated from extracts of the mollusc Anomalocardia brasiliana. The enzyme totally desulfates both chondroitin 4- and 6-sulfates without concomitant depolymerization of the compounds. It has no activity upon heparan sulfate, heparin, dermatan sulfate, and chondroitin sulfate disaccharides. It shows a pH of 5.0 and a temperature of 37 degrees C for optimum activity with a Km of 4 x 10(-5) M. The sulfatase is inhibited by sulfate and phosphate ions and HgCl2. The latter inhibition is reverted by sodium tetrathionate. Contrary to the sulfatases described so far the enzyme is activated by the lactone of D-saccharic acid when in the presence of beta-glucuronidase and beta-N-acetylgalactosaminidase. Several experiments indicate that the sulfatase is the first enzyme in the sequential degradation of chondroitin sulfate in the mollusc. This differs from the pathway of degradation of this compound in vertebrates and bacteria. 相似文献
19.
20.
Expression of chimeric genes in Caenorhabditis elegans 总被引:7,自引:0,他引:7
We have shown the expression of transformed genes in the nematode Caenorhabditis elegans using a new gene fusion system. Vectors consisting of the flanking regions of a collagen gene (col-1) or a major sperm protein gene of C. elegans fused to the Escherichia coli uidA gene, encoding beta-glucuronidase, were microinjected into worms and found to be propagated as high-copy extrachromosomal tandem arrays. We have detected beta-glucuronidase activity in transformed lines, and have shown that the activity is dependent upon the correct reading frame of the construction and on the presence of the worm sequences. The enzyme activity was shown to be encoded by the chimeric beta-glucuronidase gene by co-segregation analysis and by inactivation with specific antisera. Expression is at a very low level, and seems to be constitutive. We have used histochemical techniques to visualize the enzyme activity in embryos. 相似文献