Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.5.1.4 (deaminase)
5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The levels of pancreatic digestive enzymes, lysosomal hydrolases, and protease inhibitors were evaluated in ascites fluid from 24 patients with acute pancreatitis diagnosed as alcoholic, gallstone-induced, or idiopathic. In this group the concentrations of amylase (354 +/- 98 ng/ml), immunoreactive cationic trypsinogen (1840 +/- 238 ng/ml), and immunoreactive elastase 2 (1492 +/- 262 ng/ml) were greatly elevated in comparison to the corresponding serum values. Enzyme levels in ascites from the idiopathic pancreatitis group tended to be higher than the levels from the other two groups. Activity of acid phosphatase and beta-glucuronidase was significantly higher in ascites compared to serum in all groups. On the other hand, levels of immunoreactive alpha 1-protease inhibitor and alpha 2-macroglobulin in ascites fluid were about half the average concentrations reported for normal serum. Significant amounts of tryptic amidase activity (61.7 +/- 13.7 micrograms/ml) were observed, indicating a trypsin-alpha 2-macroglobulin complex. These data indicate an imbalance in the protease-to-inhibitor ratio in ascites fluid from patients with acute pancreatitis. Coupled with elevated ribonuclease activity (27.4 +/- 3.4 units), a positive methemalbumin test in 23 of 24 patients (1.1 +/- 0.4 mg hematin/100 ml), and an average protein concentration of 4.0 +/- 0.2 g/100 ml, these observations demonstrate that abdominal paracentesis and the biochemical analyses of ascites fluid provide useful information related to the biochemical events in acute pancreatitis and may be useful in the diagnosis of difficult cases, but their predictive value of severity remains to be established.
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PMID:Biochemical studies in peritoneal fluid from patients with acute pancreatitis. Relationship to etiology. 381 84

1. Homogenates of the mucosa of the small intestine of the guinea pig were separated by fractional sedimentation into seven different fractions. The enzymic properties of some of these subcellular fractions were compared with those obtained from the mucosa of the small intestine of the rabbit and cat. 2. The enzymic properties of the low-speed sediment (15000g-min.) were investigated and it was shown that invertase and alkaline ribonuclease were predominantly located in this subcellular fraction, whereas alkaline phosphatase, aryl-amidase, acid phosphatase, acid ribonuclease and phosphoprotein phosphatase, though true constituents of this fraction, occurred to varying degrees in other subcellular structures also. 3. It was shown that the most probable source of the enzymic activities observed in the low-speed sediment was the brush border. Electron micrographs of the purified brush-border fraction indicated vesicles derived from the brush-border membrane. 4. A method is described for the fractionation of mucosal homogenates into a brush border-plus-nuclei fraction, a mitochondrial fraction, a microsomal fraction and a particle-free supernatant. The fractions were shown to be relatively pure, as indicated by the distribution of invertase, DNA, succinate dehydrogenase, glucose 6-phosphatase and 6-phosphogluconate dehydrogenase. 5. Most of the activity of four lysosomal enzymes present in the nuclei-free homogenate was sedimented at 375000g-min., suggesting the occurrence of lysosomal particles in mucosal homogenates. 6. Further fractionation of the microsomal membranes into three fractions is described. The enzymic composition of the membrane fractions is given and discussed in relation to their structure as seen in electron micrographs.
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PMID:Studies on the fractionation of mucosal homogenates from the small intestine. 428 74

Northern blotting analysis with RNA probes derived from amidase and nitrile hydratase genes from Rhodococcus sp. ACV2 revealed that both genes are part of the same operon. RNase protection mapping and sequence analysis indicated that the operon is probably under the control of a sigma 70-like promoter located upstream from the amidase gene. Plasmids were constructed with the cloned genes under tac and lac promoter control. Expression of amdA was demonstrated in Escherichia coli. In another construction, the amdA gene was inserted under the control of the bacteriophage T7 promoter. Large amounts of recombinant amidase (at least 20% of total proteins) in a soluble and active form were obtained with the E. coli-T7 expression system by lowering the growth temperature to 29 degrees C, without IPTG induction. The ratio of amidase activity of strain ACV2 to E. coli was approximately 1:3. Purification of the recombinant amidase was carried out in one chromatographic step, giving an enzyme preparation that could be used directly in a biotechnological process.
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PMID:Transcriptional analysis of the nitrile-degrading operon from Rhodococcus sp. ACV2 and high level production of recombinant amidase with an Escherichia coli-T7 expression system. 1034 69

A major difficulty with isolating enzymatically or chemically released oligosaccharides from large-scale glycoprotein deglycosylation reactions is the time-consuming chromatography, desalting, and concentration steps required to prepare a glycan fraction of manageable proportions. To overcome these time and preparative chromatography equipment requirements, we have developed a rapid organic solvent precipitation/extraction procedure that allows sequential isolation of endo-beta-N-acetylglucosaminidase H (EC 3.2.1.96)-released high-mannose and hybrid, peptide-N(4)-(N-acetyl-beta-glucosaminyl) Asn amidase (EC 3.5.1. 52)-released complex, and beta-eliminated O-linked glycans without the need for intermediate chromatography, desalting, or concentration steps. The method involves precipitation of protein and released glycans at -20 degrees C in 80% acetone and extraction of the glycans from the pellet with 60% aqueous methanol after each deglycosylation step. Three pools of essentially salt- and detergent-free oligosaccharides (high-mannose/hybrid, complex, and O-linked) can be isolated in a high yield in 4 days with this protocol, which has been extensively tested using bovine RNase B, human bile salt-stimulated lipase expressed in Pichia pastoris, hen ovalbumin, bovine fetuin, bovine thyroglobulin, and several invertase preparations from wild-type and mutant yeast strains.
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PMID:Selective organic precipitation/extraction of released N-glycans following large-scale enzymatic deglycosylation of glycoproteins. 1066 Apr 52

Profiles of nucleotide levels in two varieties of Japanese green teas (cv. Yabukita and Saemidori), a Chinese green tea (Longjing), and two Japanese black teas (cv. Benifuuki and Benihikari) were determined and compared with that of fresh tea leaves. The concentration of 5'-nucleotides in green tea was much higher than in black tea. Nucleoside diphosphates were present in larger amounts than nucleoside triphosphates in manufactured green and black teas, whereas the triphosphates predominated in fresh tea leaves. Low levels of 3'-nucleotides were found in green and black teas. Inosine 5'-monophosphate, which is utilized as a seasoning component, was found in all manufactured teas in concentrations ranging from 50 to 200 nmol/g of dry weight. The levels of both inosine 5'-monophosphate and guanosine 5'-monophosphate were high in Chinese Longjing green tea. The unique profiles of nucleotides in manufactured teas may be a consequence of the action of degradation enzymes, such as ribonuclease, apyrase, phosphatase, nucleotidase, and adenosine 5'-monophsphate deaminase during the commercial processing of the young leaves.
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PMID:Profiles of purine and pyrimidine nucleotides in fresh and manufactured tea leaves. 1155 41

The expression of activation-induced cytidine deaminase (AID) is prerequisite to a "trifecta" of key molecular events in B cells: class-switch recombination and somatic hypermutation in humans and mice and gene conversion in chickens. Although this critically important enzyme shares common sequence motifs with apolipoprotein B mRNA-editing enzyme, and exhibits deaminase activity on free deoxycytidine in solution, it has not been shown to act on either RNA or DNA. Recent mutagenesis data in Escherichia coli suggest that AID may deaminate dC on DNA, but its putative biochemical activities on either DNA or RNA remained a mystery. Here, we show that AID catalyzes deamination of dC residues on single-stranded DNA in vitro but not on double-stranded DNA, RNA-DNA hybrids, or RNA. Remarkably, it has no measurable deaminase activity on single-stranded DNA unless pretreated with RNase to remove inhibitory RNA bound to AID. AID catalyzes dC --> dU deamination activity most avidly on double-stranded DNA substrates containing a small "transcription-like" single-stranded DNA bubble, suggesting a targeting mechanism for this enigmatic enzyme during somatic hypermutation.
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PMID:Activation-induced cytidine deaminase deaminates deoxycytidine on single-stranded DNA but requires the action of RNase. 1265 44

We have determined the crystal structures of three homologous proteins from the pathogenic protozoans Leishmania donovani, Leishmania major, and Trypanosoma cruzi. We propose that these proteins represent a new subfamily within the isochorismatase superfamily (CDD classification cd004310). Their overall fold and key active site residues are structurally homologous both to the biochemically well-characterized N-carbamoylsarcosine-amidohydrolase, a cysteine hydrolase, and to the phenazine biosynthesis protein PHZD (isochorismase), an aspartyl hydrolase. All three proteins are annotated as mitochondrial-associated ribonuclease Mar1, based on a previous characterization of the homologous protein from L. tarentolae. This would constitute a new enzymatic activity for this structural superfamily, but this is not strongly supported by the observed structures. In these protozoan proteins, the extended active site is formed by inter-subunit association within a tetramer, which implies a distinct evolutionary history and substrate specificity from the previously characterized members of the isochorismatase superfamily. The characterization of the active site is supported crystallographically by the presence of an unidentified ligand bound at the active site cysteine of the T. cruzi structure.
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PMID:Crystal structures and proposed structural/functional classification of three protozoan proteins from the isochorismatase superfamily. 1619 69

APOBEC3G (APO3G) is a cytidine deaminase that restricts replication of vif-defective human immunodeficiency virus type 1 (HIV-1). Like other members of the cellular deaminase family, APO3G has the propensity to form homo-multimers. In the current study, we investigated the functional determinants for multimerization of human APO3G and studied the role of APO3G multimerization for catalytic activity, virus encapsidation, and antiviral activity. We found that human APO3G is capable of forming multimeric complexes in transfected HeLa cells. Interestingly, multimerization of APO3G was exquisitely sensitive to RNase treatment, suggesting that interaction of APO3G subunits is facilitated or stabilized by an RNA bridge. Mutation of a conserved cysteine residue (C97) that is part of an N-terminal zinc-finger motif in APO3G abolished multimerization of APO3G; however, the C97 mutation inhibited neither in vitro deaminase activity nor antiviral function of APO3G. These results suggest that monomeric APO3G is both catalytically active and has antiviral activity. Interference studies employing either catalytically inactive or packaging-incompetent APO3G variants suggest that wild-type APO3G is packaged into HIV-1 particles in monomeric form. These results provide novel insights into the catalytic function and antiviral property of APO3G and demonstrate an important role for C97 in the RNA-dependent multimerization of this protein.
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PMID:Monomeric APOBEC3G is catalytically active and has antiviral activity. 1664 Dec 60

The deoxycytidine deaminase APOBEC3G (A3G) is expressed in human T cells and inhibits HIV-1 replication. When transfected into A3G-deficient epithelial cell lines, A3G induces catastrophic hypermutation by deaminating the HIV-1 genome. Interestingly, studies suggest that endogenous A3G in T cells induces less hypermutation than would be expected. However, to date, the specific deaminase activity of endogenous A3G in human CD4+ T cells has not been examined directly. Here, we compared deaminase activity of endogenous and exogenous A3G in various human cell lines using a standard assay and a novel, quantitative, high-throughput assay. Exogenous A3G in epithelial cell lysates displayed deaminase activity only following RNase treatment, as expected given that A3G is known to form an enzymatically inactive RNA-containing complex. Surprisingly, comparable amounts of endogenous A3G from T cell lines or from resting or activated primary CD4+ T cells exhibited minimal deaminase activity, despite RNase treatment. Specific deaminase activity of endogenous A3G in H9, CEM, and other T cell lines was up to 36-fold lower than specific activity of exogenous A3G in epithelial-derived cell lines. Furthermore, RNase-treated T cell lysates conferred a dose-dependent inhibition to epithelial cell lysates expressing enzymatically active A3G. These studies suggest that T cells, unlike epithelial-derived cell lines, express an unidentified RNase-resistant factor that inhibits A3G deaminase activity. This factor could be responsible for reduced levels of hypermutation in T cells, and its identification and blockade could offer a means for increasing antiretroviral intrinsic immunity of T cells.
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PMID:T cells contain an RNase-insensitive inhibitor of APOBEC3G deaminase activity. 1789 23

In glycoanalysis protocols, N-glycans from glycoproteins are most frequently released with peptide- N (4)-( N-acetyl-beta-glucosaminyl)asparagine amidase F (PNGase F). As the enzyme is an amidase, it cleaves the NH-CO linkage between the Asn side chain and the Asn-bound GlcNAc residue. Usually, the enzyme has a low activity, or is not active at all, on native glycoproteins. A typical example is native bovine pancreatic ribonuclease B (RNase B) with oligomannose-type N-glycans at Asn-34. However, native RNase BS, generated by subtilisin digestion of native RNase B, which comprises amino acid residues 21-124 of RNase B, is sensitive to PNGase F digestion. The same holds for carboxymethylated RNase B (RNase B (cm)). In this study, NMR spectroscopy and molecular modeling have been used to explain the differences in PNGase F activity for native RNase B, native RNase BS, and RNase B (cm). NMR analysis combined with literature data clearly indicated that the N-glycan at Asn-34 is more mobile in RNase BS than in RNase B. MD simulations showed that the region around Asn-34 in RNase B is not very flexible, whereby the alpha-helix of the amino acid residues 1-20 has a stabilizing effect. In RNase BS, the alpha-helix formed by amino acid residues 23-32 is significantly more flexible. Using these data, the possibilities for complex formation of both RNase B and RNase BS with PNGase F were studied, and a model for the RNase BS-PNGase F complex is proposed.
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PMID:The structural basis of the difference in sensitivity for PNGase F in the de-N-glycosylation of the native bovine pancreatic ribonucleases B and BS. 1829 28


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