Two dNTP triphosphohydrolases from Pseudomonas aeruginosa possess diverse substrate specificities.
Nucleotide hydrolases are known to hydrolyze not only noncanonical dNTPs to reduce the risk of mutation, but also canonical dNTPs to maintain the dNTP concentrations in the cell. dGTP triphosphohydrolase from Escherichia coli is known as an enzyme that hydrolyzes dGTP. Recently, we identified a triphosphohydrolase from Thermus thermophilus HB8 that hydrolyzes all canonical dNTPs through a complex activation mechanism. These dNTP triphosphohydrolases are widely distributed in eubacteria, but it is difficult to predict whether they possess hydrolytic activity for dGTP or dNTP. To obtain information concerning the structure-function relationships of this protein family, we characterized two dNTP triphosphohydrolases, PA1124 and PA3043, from Pseudomonas aeruginosa. Molecular phylogenic analysis showed that dNTP triphosphohydrolases can be classified into three groups. Experimentally, PA1124 had a preference for dGTP, similar to the E. coli enzyme, whereas PA3043 displayed a broad substrate specificity. Both enzymes hydrolyzed substrates in the absence of additional dNTP as an activating effector. These kinetic data suggest that PA3043 is a novel type distinct from both the E. coli and T. thermophilus enzymes. On the basis of these results, we propose that the dNTP triphosphohydrolase family should be classified into at least three subfamilies.
Mega
Ryosuke
R
Graduate School of Frontier Biological Sciences, Osaka University, Osaka, Japan.
Kondo
Naoyuki
N
Nakagawa
Noriko
N
Kuramitsu
Seiki
S
Masui
Ryoji
R
D000595:Amino Acid Sequence; D001419:Bacteria / Q000145:classification / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002384:Catalysis; D002942:Circular Dichroism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D003854:Deoxyribonucleotides / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013696:Temperature
D001426:Bacterial Proteins; D003848:Deoxyguanine Nucleotides; D003854:Deoxyribonucleotides; D010744:Phosphoric Monoester Hydrolases