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)

Concentrations of key metabolites were determined in goldfish red muscle, while muscle and blood before and after direct electrical stimulation of the myotome (60 pulses/min, amplitude 500 mV, 10 msec pulse duration, during 10 min at 20 degrees C). In white muscle, levels of ATP, aspartate and adenylate energy charge are significantly lowered while those of AMP, IMP, NH3, alpha-ketoglutarate, lactate and malate are increased. In red muscle, the only change induced by stimulation is a 160% increase of the lactate level. In white muscle, IMP-accumulation and ammonia production are equal, suggesting the AMP-deaminase reaction to be the major source of muscular ammonia. Activation of white muscle adenylosuccinate synthetase and adenylosuccinase is suggested by the conversion of aspartate into malate during increased energy demand. There is no evidence of ammonia incorporation into alanine, glutamate or glutamine.
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PMID:Goldfish muscle energy metabolism during electrical stimulation. 661 58

Porphobilinogen (PBG) deaminase catalyzes the polymerization of four PBG monopyrrole units into the linear tetrapyrrole hydroxymethylbilane necessary for the formation of chlorophyll and heme in plant cells. Degenerate oligonucleotide primers were designed based on amino acid sequence data (generated by mass spectrometry) for purified PBG deaminase from pea (Pisum sativum L.) chloroplasts. These primers were used in TaqI polymerase-catalyzed polymerase chain reaction (PCR) amplification to produce partial cDNA and nuclear genomic fragments encoding the enzyme. Subsequently, a 1.6-kb cDNA was isolated by screening a cDNA library constructed in lambda gt11 from leaf poly(A)+ RNA with the PCR products. The cDNA encodes an approximately 40-kD polypeptide containing a 46-amino acid NH2-terminal transit peptide and a mature protein of 323 amino acids. The deduced amino acid sequence of the mature pea enzyme is similar to PBG deaminases from other species and contains the conserved arginine and cysteine residues previously implicated in catalysis. Northern blot analysis indicates that the pea gene encoding PBG deaminase is expressed to varying levels in chlorophyll-containing tissues and is subject to light induction.
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PMID:Structure and expression of chloroplast-localized porphobilinogen deaminase from pea (Pisum sativum L.) isolated by redundant polymerase chain reaction. 751 80

Candida albicans and other pathogenic Candida species can use N-acetylglucosamine as a sole carbon source for growth. GlcNAc induces the enzymes of GlcNAc catabolic pathway; besides, under certain conditions, GlcNAc also induces a change from the yeast to germ tube morphology. Glucosamine-6-phosphate deaminase (EC 5.3.1.10) is the terminal enzyme of the GlcNAc catabolic pathway. We have purified the deaminase from C. albicans and studied its characteristics. The size of the deaminase estimated from SDS-polyacrylamide gel electrophoresis is 28 kDa. N-Acetylglucosamine 6-phosphate, an allosteric activator of the Escherichia coli deaminase, has no effect on the activity of the C. albicans enzyme. The deaminase is induced over 100-fold by GlcNAc and its level is about 0.3-0.5% of the proteins in crude extract. Three cDNA clones were obtained from a lambda gt11 expression library by immunoscreening with deaminase antiserum. C. albicans genomic DNA blot hybridization revealed that the NAG1 gene, encoding the glucosamine-6-phosphate deaminase, is present in a single copy. Hybrid-selected translation and immunoprecipitation experiments revealed that the purified deaminase and the protein encoded by the clones were similar in size and in their antigenicity. DNA sequencing revealed that the largest cDNA clone contained the complete open reading frame, which can code for a 27.5-kDa protein. The NH2-terminal sequence (35 residues) determined from the purified deaminase was identical to the sequence of the deduced protein. The Nag1 protein has about 47% identity with the sequence of the E. coli glucosamine-6-phosphate deaminase. Furthermore, RNA blot hybridization showed that GlcNAc induces the expression of NAG1 gene.
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PMID:Molecular cloning and analysis of the NAG1 cDNA coding for glucosamine-6-phosphate deaminase from Candida albicans. 768 45

The combination method of carboxypeptidase Y digestion and fast atom bombardment (FAB) mass spectrometry is described for the identification of C-terminal amino acid amides in peptides. Carboxypeptidase Y has amidase activity as well as exopeptidase activity in the same digestion buffer condition. Based on this concept, we develop a new technique which can definitively and easily identify the C-terminal amino acid amides. This method obviates the need for several complicated steps occurring in previous methods, but improves sensitivity, and enables exact identification of the amino acid amide by the difference of molecular mass. Analyses of carboxypeptidase Y digested peptides, not liberated free amino acid amides, were carried out by fast atom bombardment mass spectrometry. The use of truncated peptides by fast atom bombardment mass spectrometry in C-terminal amino acid amide determination gives several advantages over analyses of the liberated amino acid amides. The C-terminal amino acid amides of Allantostatin I (Leu-NH2), alpha-Melanocyte Stimulating Hormone (Val-NH2), and Ranatensin (Met-NH2) are unequivocally determined at a level of 0.90-2.3 nmol per peptide. This approach is based on entirely different principles than the previous approaches.
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PMID:Identification of the C-terminal amino acid amides by carboxypeptidase Y digestion and fast atom bombardment mass spectrometry. 770 6

Co- and post-translational amino-terminal processing of proteins is one mechanism by which intracellular proteins can be either protected from or targeted to degradation by the N-end Rule pathway (Bachmair, A., Finley, D., and Varshavsky, A. (1986) Science 234, 179-186). A novel enzyme, protein NH2-terminal asparagine amidohydrolase, which can function in this pathway by potentially directing critical regulatory proteins possessing an amino-terminal asparagine residue formed from the removal of N-acetylmethionine, has recently been purified and characterized (Stewart, A.E., Arfin, S. M., and Bradshaw, R. A. (1994) J. Biol. Chem. 269, 23509-23517). Here, we report the isolation and characterization of a cDNA for porcine protein NH2-terminal asparagine amidohydrolase, which indicates that it is a new type of enzyme, not homologous to any previously identified protein. This provides strong evidence for the importance of regulated protein degradation in cellular functioning.
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PMID:The sequence of porcine protein NH2-terminal asparagine amidohydrolase. A new component of the N-end Rule pathway. 781 82

A constitutively expressed aliphatic amidase from a Rhodococcus sp. catalyzing acrylamide deamination was purified to electrophoretic homogeneity. The molecular weight of the native enzyme was estimated to be 360,000. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the purified preparation yielded a homogeneous protein band having an apparent molecular weight of about 44,500. The amidase had pH and temperature optima of 8.5 and 40 degrees C, respectively, and its isoelectric point was pH 4.0. The amidase had apparent K(m) values of 1.2, 2.6, 3.0, 2.7, and 5.0 mM for acrylamide, acetamide, butyramide, propionamide, and isobutyramide, respectively. Inductively coupled plasma-atomic emission spectometry analysis indicated that the enzyme contains 8 mol of iron per mol of the native enzyme. No labile sulfide was detected. The amidase activity was enhanced by, but not dependent on Fe(2+), Ba(2+), and Cr(2+). However, the enzyme activity was partially inhibited by Mg(2+) and totally inhibited in the presence of Ni(2+), Hg(2+), Cu(2+), Co(2+), specific iron chelators, and thiol blocking reagents. The NH2-terminal sequence of the first 18 amino acids displayed 88% homology to the aliphatic amidase of Brevibacterium sp. strain R312.
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PMID:Purification and characterization of an amidase from an acrylamide-degrading Rhodococcus sp. 794 67

Porphobilinogen deaminase (EC 4.3.1.8) has been purified to homogeneity (16,000-fold) from the plant Arabidopsis thaliana in yields of 8%. The deaminase is a monomer of M(r) 35,000, as shown by SDS/PAGE, and 31,000, using gel-filtration chromatography. The pure enzyme has a Vmax. of 4.5 mumol/h per mg and a Km of 17 +/- 4 microM. Determination of the pI and pH optimum revealed values of 5.2 and 8.0 respectively. The sequence of the N-terminus was found to be NH2-XVAVEQKTRTAI. The deaminase is heat-stable up to 70 degrees C and is inhibited by NH3 and hydroxylamine. The enzyme is inactivated by arginine-, histidine- and lysine-specific reagents. Incubation with the substrate analogue and suicide inhibitor, 2-bromoporphobilinogen, results in chain termination and in inactivation.
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PMID:Purification and properties of porphobilinogen deaminase from Arabidopsis thaliana. 819 81

Adenosine 5'-monophosphate (AMP) deaminase from baker's yeast is an allosteric enzyme containing a single AMP binding site and two ATP regulatory sites per polypeptide [Merkler, D. J., & Schramm, V. L. (1990) J. Biol Chem. 265, 4420-4426]. The enzyme contains 0.98 +/- 0.17 zinc atom per subunit. The X-ray crystal structure for mouse adenosine deaminase shows zinc in contact with the attacking water nucleophile using purine riboside as a transition-state inhibitor [Wilson, D. K., Rudolph, F. B., & Quiocho, F. A. (1991) Science 252, 1278-1284]. Alignment of the amino acid sequence for yeast AMP deaminase with that for mouse adenosine deaminase demonstrates conservation of the amino acids known from the X-ray crystal structure to bind to the zinc and to a transition-state analogue. On the basis of these similarities, yeast AMP deaminase is also proposed to use a Zn(2+)-activated water molecule to attack C6 of AMP with the displacement of NH3. The pKm and pKi profiles for AMP and a competitive inhibitor overlap in a bell-shaped curve with pKa values of 7.0 and 7.4. This pattern is characteristic of a rapid equilibrium between AMP and the enzyme, thus confirming the rapid equilibrium random kinetic patterns [Merkler, D. J., Wali, A. S., Taylor, J., Schramm, V. L. (1989) J. Biol. Chem. 264, 21422-21430]. The Vmax of the reaction requires one unprotonated and one protonated group with pKa values of 6.4 +/- 0.2 and 7.7 +/- 0.3, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Catalytic mechanism of yeast adenosine 5'-monophosphate deaminase. Zinc content, substrate specificity, pH studies, and solvent isotope effects. 850 99

Escherichia coli asparagine synthetase B (AS-B) catalyzes the synthesis of asparagine from aspartic acid and glutamine in an ATP-dependent reaction. The ability of this enzyme to employ hydroxylamine and L-glutamic acid gamma-monohydroxamate (LGH) as alternative substrates in place of ammonia and L-glutamine, respectively, has been investigated. The enzyme is able to function as an amidohydrolase, liberating hydroxylamine from LGH with high catalytic efficiency, as measured by k(cat)/K(M). In addition, the kinetic parameters determined for hydroxylamine in AS-B synthetase activity are very similar to those of ammonia. Nitrogen transfer from LGH to yield aspartic acid beta-monohydroxamate is also catalyzed by AS-B. While such an observation has been made for a few members of the trpG amidotransferase family, our results appear to be the first demonstration that nitrogen transfer can occur from glutamine analogs in a purF amidotransferase. However, k(cat)/K(M) for the ATP-dependent transfer of hydroxylamine from LGH to aspartic acid is reduced 3-fold relative to that for glutamine-dependent asparagine synthesis. Further, the AS-B mutant in which asparagine is replaced by alanine (N74A) can also use hydroxylamine as an alternate substrate to ammonia and catalyze the hydrolysis of LGH. The catalytic efficiencies (k(cat)/K(M)) of nitrogen transfer from LGH and L-glutamine to beta-aspartyl-AMP are almost identical for the N74A AS-B mutant. These observations support the proposal that Asn-74 plays a role in catalyzing glutamine-dependent nitrogen transfer. We interpret our kinetic data as further evidence against ammonia-mediated nitrogen transfer from glutamine in the purF amidotransferase AS-B. These results are consistent with two alternate chemical mechanisms that have been proposed for this reaction [Boehlein, S. K., Richards, N. G. J., Walworth, E. S., & Schuster, S. M. (1994) J. Biol. Chem. 269, 26789-26795].
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PMID:Glutamic acid gamma-monohydroxamate and hydroxylamine are alternate substrates for Escherichia coli asparagine synthetase B. 860 42

LytA amidase is the best known bacterial autolysin. It breaks down the N-acetylmuramoyl-L-alanine bonds in the peptidoglycan backbone of Streptococcus pneumoniae and requires the presence of choline residues in the cell-wall teichoic acids for activity. Genetic experiments have supported the hypothesis that its 36-kDa chain has evolved by the fusion of two independent modules: the NH2-terminal module, responsible for the catalytic activity, and the COOH-terminal module, involved in the attachment to the cell wall. The structural organization of LytA amidase and of its isolated COOH-terminal module (C-LytA) and the variations induced by choline binding have been examined by differential scanning calorimetry and analytical ultracentrifugation. Deconvolution of calorimetric curves have revealed a folding of the polypeptide chain in several independent or quasi-independent cooperative domains. Elementary transitions in C-LytA are close but not identical to those assigned to the COOH-terminal module in the complete amidase, particularly in the absence of choline. These results indicate that the NH2-terminal region of the protein is important for attaining the native tertiary fold of the COOH terminus. Analytical ultracentrifugation studies have shown that LytA exhibits a monomer <--> dimer association equilibrium, through the COOH-terminal part of the molecule. Dimerization is regulated by choline interaction and involves the preferential binding of two molecules of choline per dimer. Sedimentation velocity experiments give frictional ratios of 1.1 for C-LytA monomer and 1.4 for C-LytA and LytA dimers; values that deviated from that of globular rigid particles. When considered together, present results give evidence that LytA amidase might be described as an elongated molecule consisting of at least four domains per subunit (two per module) designated here in as N1, N2, C1, and C2. Intersubunit cooperative interactions through the C2 domain in LytA dimer occur under all experimental conditions, while C-LytA requires the saturation of low affinity choline binding sites. The relevance of the structural features deduced here for LytA amidase is examined in connection with its biological function.
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PMID:Structural organization of the major autolysin from Streptococcus pneumoniae. 863 7


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