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)

A 138-kDa glycoprotein comprising folate deaminase activity was purified to apparent homogeneity from membranes of Dictyostelium discoideum. Deaminase activity could be effectively inhibited by p-chloromercuriphenylsulfonate. This treatment protected folate from deamination and thus allowed investigation of folate binding to deaminase fractions. Two types of folate binding sites, differing in affinity and specificity, were detected on the folate deaminase glycoprotein. One type displays high affinity and binds folate stronger than N10-methylfolate. This binding site appears to be identical with the catalytic site of folate deaminase. The other type of binding site shows lower affinity but prefers N10-methylfolate relative to folate. A similar preference for N10-methylfolate was observed in chemotaxis tests pointing to the possibility that the second type of binding site is involved in chemotactic perception of folate compounds. Folate perception and deamination could thus be performed by activities residing on the same polypeptide.
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PMID:A 138-kDa glycoprotein from Dictyostelium membranes with folate deaminase and folate binding activity. 154 93

A method has been developed for the efficient selection of chemotaxis mutants of Dictyostelium discoideum. Mutants defective in the chemotactic response to folate could be enriched up to 30-fold in one round of selection using a chamber in which a compartment that contained the chemoattractant was separated by a sandwich of four nitrocellulose filters from a compartment that contained buffer. Mutagenized cells were placed in the center of the filter layer and exposed to the attractant gradient built up between the compartments for a period of 3-4 h. While wild-type cells moved through the filters in a wave towards the compartment that contained attractant, mutant cells remained in the filter to which they were applied. After several repetitions of the selection procedure, mutants defective in chemotaxis made up 10% of the total cell population retained in that filter. Mutants exhibiting three types of alterations were collected: motility mutants with either reduced speed of movement, or altered rates of turning; a single mutant defective in production of the attractant-degrading enzyme, folate deaminase; and mutants with normal motility but reduced chemotactic responsiveness. One mutant showed drastically reduced sensitivity in folate-induced cGMP production. Morphogenetic alterations of mutants defective in folate chemotaxis are described.
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PMID:Selection of chemotaxis mutants of Dictyostelium discoideum. 379 59

Cyclic AMP and folic acid act as chemotactic factors in Dictyostelium discoideum. Both agents, when applied extracellularly, also control cell development from the growth stage to the acquisition of aggregation competence. Cyclic AMP phosphodiesterase and folate deaminase are extracellular enzymes whose activity is regulated during early differentiation of D. discoideum cells. The two enzymes help control the extracellular levels of cyclic AMP and folic acid. The substrates cyclic AMP and folic acid each increase the extracellular activity of folate deaminase as well as phosphodiesterase. The specificity of extracellular phosphodiesterase regulation by cyclic AMP indicates that the effect is mediated by specific cyclic AMP receptors rather than the catalytic site of cell surface phosphodiesterase. To some extent cyclic AMP and folic acid are interchangeable with respect to regulating differentiation and enhancing enzymatic inactivation of intercellular signals. Thus the two extracellular signals may share a common cellular pathway of signal transduction. The regulation of folate deaminase and phosphodiesterase by folic acid does not always parallel the folic acid effects on development. Pulses of folic acid stimulate development of aggregation competence, whereas a continuous flux inhibits. In contrast, either continuous flux or pulses of folic acid increase the deaminase and phosphodiesterase activities.
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PMID:Folate deaminase and cyclic AMP phosphodiesterase in Dictyostelium discoideum: their regulation by extracellular cyclic AMP and folic acid. 626 63

Kinetic data obtained for deamination of pterin by the extracellular fraction from Dictyostelium discoideum yielded apparently linear Lineweaver-Burk plots for pterin. The Michaelis constant for pterin was 30 microM. The data for folic acid deamination yielded convex Lineweaver-Burk plots. Convex Lineweaver-Burk plots could result from the presence of two types of enzymes with different affinities. The data for folic acid deamination were analyzed mathematically for two types of enzymes. This analysis produced Michaelis constants for folic acid of 1.8 and 23 microM competition studies suggested that an enzyme with low affinity nonspecifically catalyzed the deamination of folic acid and pterin, whereas an enzyme with high affinity was a specific folic acid deaminase. A specific folic acid deaminase with high affinity appeared to be present on the surface of D. discoideum cells. The Michaelis constant for this enzyme was 2.6 microM. Cells growing in nutrient broth and cells starved in phosphate buffer released folic acid and pterin deaminases. The quantity of deaminase activities released by the cells appeared to be controlled by chemoattractants. Starving cells that were supplied with folic acid, pterin, or adenosine 3',5'-phosphate increased their extracellular folic acid and pterin deaminase activities to a larger extent than did cell suspensions to which no chemoattractants were added. Administration of folic acid or pterin to starving cells caused increases of the activity of extracellular adenosine 3',5'-phosphate phosphodiesterase and repressed increases of the activity of phosphodiesterase inhibitor.
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PMID:Folic acid and pterin deaminases in Dictyostelium discoideum: kinetic properties and regulation by folic acid, pterin, and adenosine 3',5'-phosphate. 627 62

Folate deaminase released from cells of Dictyostelium discoideum is heterogeneous with respect to molecular weight and stability at 60 degrees C. The most heat-stable component isoelectrofocuses in a broad band at approx. pH 6. The Km value of this component for folate is approx. 7 x 10(-7)M and Mr approx. 40 000. The major portion if not all of the deaminase binds to immobilized concanavalin A and lentil lectin. Extracellular folate deaminase has a pH-optimum of approx. pH 6.0. This is higher than that of lysosomal enzymes, which are also glycoproteins released into the extracellular medium.
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PMID:Extracellular folate deaminase of Dictyostelium discoideum. 627 Dec 53

Studies of the folate chemotactic receptor of vegetative Dictyostelium discoideum cells have been hampered by the presence of the degradative enzyme folate deaminase. The diaminopterin compounds aminopterin and methotrexate (MTX) are chemoattractants but are not attacked by the deaminase. [3',5',7,9-3H]methotrexate ([3H]MTX) is a nondegraded radioligand for the folate receptor. Binding to the receptor is rapid, reaching steady state in less than one min, and reversible in less than 15 s by an excess of unlabeled MTX. A single class of binding sites is found with a Kd of 2 x 10(-8) M, which correlates well with the concentration dependence of chemotaxis. Folate, aminopterin, and MTX all compete for [3H]MTX binding, whereas pterin, p-aminobenzoate, and nucleotides do not. Analysis of the receptor during differentiation indicates a decrease in site number by a factor of 3 with no change in affinity during the first 7 hr. During this time, the directional response (chemotaxis) to MTX and folate is lost, but a nondirectional stimulation of motility rate (chemokinesis) is retained. The response to cyclic AMP displays reciprocal behavior, first appearing as a chemokinetic response and then as a chemotactic response.
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PMID:[3H]Methotrexate as a ligand for the folate receptor of Dictyostelium discoideum. 627 68

The use of high-performance liquid chromatography to identify and quantitate five purine-metabolizing enzymes from a partially purified subcellular fraction of the eucaryotic microorganism Dictyostelium discoideum is described. All HPLC separations were carried out in an isocratic manner using reverse-phase C18 as the stationary phase. The mobile phase consisted of a phosphate buffer with either methanol or acetonitrile as cosolvent, and optimal separation conditions were attained by varying the organic concentration or the pH of the buffer or by employing paired-ion chromatographic techniques. Substrates and products were detected at either 254 nm for the purines or 295 nm for the formycin analogs. An adenosine kinase activity was identified, and it was demonstrated that formycin A (FoA) could be substituted for adenosine as the phosphate acceptor, yielding FoAMP as the product. With FoA as the substrate an apparent Km of 18.2 microM and an apparent Vmax of 32.4 mmol min-1 mg-1 were observed for the activity. A purine-nucleoside phosphorylase activity was found to cleave adenosine to adenine and ribosylphosphate. FoA was not found to be a substrate for this activity due to the unusual formycin C-glycosyl bond which was not hydrolyzed by enzymes or chemically with either HCl or NaOH. An adenylate deaminase activity was found to be present in the cytosolic S-100 of cells harvested during the onset of development, and this deaminase activity was greatly stimulated by ATP. With FoAMP as the substrate, an apparent Km of 236 microM and Vmax of 2.78 mumol min-1 mg-1 were observed. The deamination of FoAMP could be inhibited by the addition of the natural substrate AMP. An apparent Ki value of 136 microM was determined from initial rate data. An adenylosuccinate synthetase activity was observed to have a Km value for GTP, IMP, and aspartic acid of 23, 34, and 714 microM, respectively. The formycin analog FoIMP was not a substrate with this activity but was a competitive inhibitor of IMP. Finally hypoxanthine-guanine phosphoribosyltransferase was found to have Km and Vmax values for hypoxanthine of 55.5 microM and 34.3 nmol-1 min-1 mg-1. When guanine was used as the substrate, the rate of nucleotide formation was 50% that with hypoxanthine as the substrate. The advantages of using HPLC to examine the interconnecting activities of a multienzyme complex in subcellular fractions are discussed, including the increased sensitivity obtained by using formycin analogs in the assay procedures.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Intermediary purine-metabolizing enzymes from the cytosol of Dictyostelium discoideum monitored by high-performance liquid chromatography. 642 68

Two specific fatty acyl amidases that hydrolyze lipopolysaccharide have been isolated from the slime mold Dictyostelium discoideum. Esterases as well as phosphatases acting on lipid A derivatives were also observed. The first amidase (I) hydrolyzes the fatty amide adjacent to the C-1 phosphate on the disaccharide backbone of lipid A. Amidase II cleaves the distal amide, but only after deacylation of the first site. The range of specificity and the structural determinants important to specificity of the amidases were evaluated in studies of specifically modified derivatives of lipid A. In light of the effects of lipopolysaccharide on the biology of D. discoideum, a role for the amidases and other lipopolysaccharide-specific catabolic enzymes is discussed.
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PMID:Lipases specifically degrading lipopolysaccharide: fatty acyl amidases from Dictyostelium discoideum. 647 5

Two amidases have been partially purified from the slime mold Dictyostelium discoideum; these act sequentially on the beta-hydroxymyristyl-amide groups present in the lipopolysaccharide derivative (4'-O-phosphoryl-N-beta-hydroxymyristyl-D-glucosaminyl)-beta-(1 leads to 6)-N-beta-hydroxymyristyl-D-glucosamine-1-phosphate (III). Amidase-I, which specifically removes the myristyl chain near the 1-phosphate of compound III (apparent Km, 3.7 microM), has been purified 110-fold from a lysate of D. discoideum NC4 cultivated on Escherichia coli. The partially purified enzyme contains no other amidase or phosphatase activities; however, an esterase activity can be detected. The second amidase has been purified about 12-fold from the extracellular fluid of D. discoideum AX3 cultured axenically. This amidase hydrolyzes the distal amide linkage in III (apparent Km, approximately 20 microM) only after prior deacylation of the first site by amidase-I. The preparation is free from phosphatases and glycosidases that can act on lipopolysaccharide. The differential expression of the amidases in D. discoideum and some of their kinetic properties have been described. The amidases should prove useful in structure-function studies of lipopolysaccharide.
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PMID:Fatty acyl amidases from Dictyostelium discoideum that act on lipopolysaccharide and derivatives. I. Partial purification and properties. 710 2

The substrate specificities of two fatty acyl amidases partially purified from the slime mold Dictyostelium discoideum have been studied. The amidase act on lipopolysaccharide derivatives, such as (4'-O-phosphoryl-N-beta-hydroxymyristyl-D-glucosaminyl)-beta-(1 leads to 6)-N-beta-hydroxymyristyl-D-glucosamine-1-phosphate (III) in a sequential manner. Amidase-I removes the beta-hydroxymyristyl residue present on the amino group adjacent to the 1-phosphate and the product formed is a substrate for amidase-II; the latter removes the remaining beta-hydroxymyristyl residue from the distal amino group. Compound III itself is resistant to amidase-II. Removal of the C-1 or C-4 phosphate groups does not influence recognition by the amidases or their sequential action. Both amidases are specific for long chain fatty amide linkages. Thus, a formyl group on the glucosamine amino group adjacent to the C-1 phosphate is not hydrolyzed by amidase-I; however, this substituent does not hinder the action of amidase-II on the distal fatty acyl amide. The presence of the beta-hydroxyl group in myristyl-amide residues is not required for hydrolysis. Further, while amidase-I requires disaccharide structures for its action, amidase-II acts on monosaccharides as well. Finally, the effects of a variety of substrate analogs and divalent ions on the activity of the enzymes are reported.
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PMID:Fatty acyl amidases from Dictyostelium discoideum that act on lipopolysaccharide and derivatives. II. Aspects of substrate specificity. 710 3


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