Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.5.4.4 (adenosine deaminase)
 5,136 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Murine adenosine deaminase (mADA) is a 40 kDa (beta/alpha)(8)-barrel protein consisting of eight central beta-strands and eight peripheral alpha-helices containing four tryptophan residues. In this study, we investigated the urea-dependent behavior of the protein labeled with 6-fluorotryptophan (6-(19)F-Trp). The (19)F NMR spectrum of 6-(19)F-Trp-labeled mADA reveals four distinct resonances in the native state and three partly overlapped resonances in the unfolded state. The resonances were assigned unambiguously by site-directed mutagenesis. Equilibrium unfolding of 6-(19)F-Trp-labeled mADA was monitored using (19)F NMR based on these assignments. The changes in intensity of folded and unfolded resonances as a function of urea concentration show transition midpoints consistent with data observed by far-UV CD and fluorescence spectroscopy, indicating that conformational changes in mADA during urea unfolding can be followed by (19)F NMR. Chemical shifts of the (19)F resonances exhibited different changes between 1.0 and 6.0 M urea, indicating that local structures around 6-(19)F-Trp residues change differently. The urea-induced changes in local structure around four 6-(19)F-Trp residues of mADA were analyzed on the basis of the tertiary structure and chemical shifts of folded resonances. The results reveal that different local regions in mADA have different urea-dependent behavior, and that local regions of mADA change sequentially from native to intermediate topologies on the unfolding pathway.
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PMID:Urea-dependent unfolding of murine adenosine deaminase: sequential destabilization as measured by 19F NMR. 1476 19

Adenosine deaminase (ADA, EC 3.5.4.4) is a ubiquitous (beta/alpha)8-barrel enzyme crucial for purine metabolism and normal immune competence. In this study, it was observed that loss of enzyme activity of murine ADA (mADA) precedes the global secondary and tertiary structure transition when the protein is exposed to denaturant. The structural mechanism for this phenomenon was probed using site-specific 19F NMR spectroscopy in combination with [6-19F]tryptophan labeling and inhibitor binding. There are four tryptophan residues in mADA and all are located more than 12 A from the catalytic site. The 19F NMR spectra of [6-19F]Trp-labelled mADA show that the urea-induced chemical shift change of 19F resonance of W161, one of the four tryptophan 19F nuclei, correlates with the loss of enzyme activity. The urea-induced chemical shift change of another 19F resonance of W117 correlates with the change of the apparent rate constant for the binding of transition-state analogue inhibitor deoxycoformycin to the enzyme. On the other hand, the chemical environment of the local region around W264 does not change significantly, as a consequence of perturbation by low concentrations of urea or substrate analog. The results indicate that different regions of mADA have different local stability, which controls the activity and stability of the enzyme. The results provide new insights into the relationship between the function of a protein and its conformational flexibility as well as its global stability. This study illustrates the advantage of 19F NMR spectroscopy in probing site-related conformational change information in ligand binding, enzymatic activity and protein folding.
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PMID:Relation of enzyme activity to local/global stability of murine adenosine deaminase: 19F NMR studies. 1558 1

Alpha-amino-beta-carboxymuconate-epsilon-semialdehyde decarboxylase (ACMSD) is a widespread enzyme found in many bacterial species and all currently sequenced eukaryotic organisms. It occupies a key position at the branching point of two metabolic pathways: the tryptophan to quinolinate pathway and the bacterial 2-nitrobenzoic acid degradation pathway. The activity of ACMSD determines whether the metabolites in both pathways are converted to quinolinic acid for NAD biosynthesis or to acetyl-CoA for the citric acid cycle. Here we report the first high-resolution crystal structure of ACMSD from Pseudomonas fluorescens which validates our previous predictions that this enzyme is a member of the metal-dependent amidohydrolase superfamily of the (beta/alpha)(8) TIM barrel fold. The structure of the enzyme in its native form, determined at 1.65 A resolution, reveals the precise spatial arrangement of the active site metal center and identifies a potential substrate-binding pocket. The identity of the native active site metal was determined to be Zn. Also determined was the structure of the enzyme complexed with cobalt at 2.50 A resolution. The hydrogen bonding network around the metal center suggests that Arg51 and His228 may play important roles in catalysis. The metal center configuration of PfACMSD is very similar to that of Zn-dependent adenosine deaminase and Fe-dependent cytosine deaminase, suggesting that ACMSD may share certain similarities in its catalytic mechanism with these enzymes. These data enable us to propose possible catalytic mechanisms for ACMSD which appear to be unprecedented among all currently characterized decarboxylases.
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PMID:Crystal structure of alpha-amino-beta-carboxymuconate-epsilon-semialdehyde decarboxylase: insight into the active site and catalytic mechanism of a novel decarboxylation reaction. 1693 94

The effect of inhibitors, 1-deazaadenosine (1-dAdo) and erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), on the conformation of adenosine deaminase was studied using the method of selective quenching of fluorescence emission by acrylamide, I- and Cs+. Both in free adenosine deaminase and in its complexes with the inhibitors, the wavelength maxima and half-width of the emission characterize the environment of fluorescing tryptophan residues in adenosine deaminase as weak polar with limited access to solvent. The formation of complexes with the ground state inhibitors used did not quench or change the main emission characteristics of tryptophan fluorescence in adenosine deaminase. Small blue shifts of emission maxima were observed upon quenching in all three samples. The Stern-Volmer parameters of tryptophan fluorescence quenching by acrylamide were not essentially influenced by complex formation of the enzyme with the inhibitors: in general, the folding of the enzyme molecule in the complexes is not perturbed. On the contrary, the emission quenching by charged heavy ions, I- and Cs+, in the complexes was hindered in comparison with free adenosine deaminase. In the complex with 1-deazaadenosine, the parameters for quenching by both ions evidence the essential worsening of their interaction with tryptophans. In the complex with erythro-9-(2-hydroxy-3-nonyl)adenine, along with the worse quenching by I-, complete prohibition of quenching by Cs+ was observed. These data indicate that the local environments of fluorescing tryptophan residues is substantially distorted compared with free adenosine deaminase, which leads to their screening from charged heavy ions.
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PMID:[Conformation of adenosine deaminase in complexes with inhibitors: application of selective quenching of fluorescence emission]. 1854 63

A metabolomic analysis of plasma amino acids and acylcarnitines was applied to four disorders of nucleotide metabolism. Multivariate analysis gave score plots that show segregation of hypoxanthine phosphoribosyltransferase and adenine phosphoribosyltransferase deficient plasma from controls with equivocal results for adenosine deaminase and dihydropyrimidine dehydrogenase deficiencies. Loadings plots revealed the principal metabolites responsible for the discrimination between these classes. There were increases for HPRT in C4-, C6-, and C3-DC (malonyl)-carnitines, and decreased serine. For APRT there were increases in C4- to C10- and C3-DC to C6-DC-carnitines, urea, 1-methylhistidine, 3-methylhistidine, and decreased tryptophan. For ADA deficiency there were increases in C4- and C6-carnitines, taurine, and isoleucine.
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PMID:Application of metabolomic principles to disorders of nucleotide metabolism reveals new metabolic perturbations. 1860 May 20

Severe combined immunodeficiencies (SCIDs) appear as optimal disease targets to challenge potential efficacy of gene therapy. Ex vivo, retrovirally mediated gene transfer into hematopoietic progenitor cells has been shown to provide sustained correction of two forms of SCID, that is, SCID-X1 and adenosine deaminase deficiencies. In the former case, however, genotoxicity was observed in a minority of patients as a consequence of retroviral integration into proto-oncogenes loci and transactivation. Design of vectors in which the enhancer element of retroviral LTR has been deleted and an internal promoter added (self-inactivated vectors) could provide both safe and efficient gene transfer as being presently tested.
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PMID:Strategies for retrovirus-based correction of severe, combined immunodeficiency (SCID). 2236 67


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