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

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

Proteins are intrinsically flexible macromolecules that undergo internal motions with time scales spanning femtoseconds to milliseconds. These fluctuations are implicated in the optimization of reaction barriers for enzyme catalyzed reactions. Time, temperature, and mutation dependent hydrogen-deuterium exchange coupled to mass spectrometry (HDX-MS) has been previously employed to identify spatially resolved, catalysis-linked dynamical regions of enzymes. We now extend this technique to pursue the correlation of protein flexibility and chemical reactivity within the diverse and widespread TIM barrel proteins, targeting murine adenosine deaminase (mADA) that catalyzes the irreversible deamination of adenosine to inosine and ammonia. Following a structure-function analysis of rate and activation energy for a series of mutations at a second sphere phenylalanine positioned in proximity to the bound substrate, the catalytically impaired Phe61Ala with an elevated activation energy (Ea = 7.5 kcal/mol) and the wild type (WT) mADA (Ea = 5.0 kcal/mol) were selected for HDX-MS experiments. The rate constants and activation energies of HDX for peptide segments are quantified and used to assess mutation-dependent changes in local and distal motions. Analyses reveal that approximately 50% of the protein sequence of Phe61Ala displays significant changes in the temperature dependence of HDX behaviors, with the dominant change being an increase in protein flexibility. Utilizing Phe61Ile, which displays the same activation energy for kcat as WT, as a control, we were able to further refine the HDX analysis, highlighting the regions of mADA that are altered in a functionally relevant manner. A map is constructed that illustrates the regions of protein that are proposed to be essential for the thermal optimization of active site configurations that dominate reaction barrier crossings in the native enzyme.
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PMID:Hydrogen-Deuterium Exchange within Adenosine Deaminase, a TIM Barrel Hydrolase, Identifies Networks for Thermal Activation of Catalysis. 3318 Oct 18