Author(s):
Rodrigues, Joaquim Rui ; Fernández, Ascensión ; Canales, José ; Cabezas, Alicia ; Ribeiro, João Meireles ; Costas, María Jesús ; Cameselle, José Carlos
Date: 2012
Persistent ID: http://hdl.handle.net/10400.8/3008
Origin: IC-online
Subject(s): Adenosine Diphosphate Ribose; Animals; Binding Sites; Catalytic Domain; Cyclic AMP; Cytidine Diphosphate; Enzyme Activation; Hydrogen-Ion Concentration; Magnesium; Manganese; Molecular Docking Simulation; Mutation; Pyrophosphatases; Rats; Substrate Specificity; Zebrafish Proteins; Zebrafish; Adenosine Diphosphate Ribose; Adenosine Diphosphate Ribose; Animals; Animals; Binding Sites; Binding Sites; Catalytic Domain; Catalytic Domain; Cyclic AMP; Cyclic AMP; Cytidine Diphosphate; Cytidine Diphosphate; Enzyme Activation; Enzyme Activation; Hydrogen-Ion Concentration; Hydrogen-Ion Concentration; Magnesium; Magnesium; Manganese; Manganese; Molecular Docking Simulation; Molecular Docking Simulation; Mutation; Mutation; Pyrophosphatases; Pyrophosphatases; Rats; Rats; Substrate Specificity; Substrate Specificity; Zebrafish Proteins; Zebrafish Proteins; Zebrafish; Zebrafish
Description
The ADPRibase-Mn-like protein family, that belongs to the metallo-dependent phosphatase superfamily, has different functional and structural prototypes. The functional one is the Mn(2+)-dependent ADP-ribose/CDP-alcohol diphosphatase from Rattus norvegicus, which is essentially inactive with Mg(2+) and active with low micromolar Mn(2+) in the hydrolysis of the phosphoanhydride linkages of ADP-ribose, CDP-alcohols and cyclic ADP-ribose (cADPR) in order of decreasing efficiency. The structural prototype of the family is a Danio rerio protein with a known crystallographic structure but functionally uncharacterized. To estimate the structure-function correlation with the same protein, the activities of zebrafish ADPRibase-Mn were studied. Differences between zebrafish and rat enzymes are highlighted. The former showed a complex activity dependence on Mn(2+), significant (≈25%) Mg(2+)-dependent activity, but was almost inactive on cADPR (150-fold less efficient than the rat counterpart). The low cADPR hydrolase activity agreed with the zebrafish genome lacking genes coding for proteins with significant homology with cADPR-forming enzymes. Substrate-docking to zebrafish wild-type protein, and characterization of the ADPRibase-Mn H97A mutant pointed to a role of His-97 in catalysis by orientation, and to a bidentate water bridging the dinuclear metal center as the potential nucleophile. Finally, three structural elements that delimit the active site entrance in the zebrafish protein were identified as unique to the ADPRibase-Mn-like family within the metallo-dependent phosphatase superfamily.
