Characterization of the Mn2+-stimulated (di)adenosine polyphosphate hydrolase encoded by the Deinococcus radiodurans DR2356 nudix gene.
The DR2356 nudix hydrolase gene from Deinococcus radiodurans has been cloned and the product expressed as an 18 kDa histidine-tagged protein. The enzyme hydrolysed adenosine and diadenosine polyphosphates, always generating ATP as one of the initial products. ATP and other (deoxy)nucleoside triphosphates were also substrates, yielding (d)NDP and Pi as products. The DR2356 protein was most active at pH 8.6-9.0 and showed a strong preference for Mn(2+) as activating cation. Mg(2+) ions at 15 mM supported only 5% of the activity achieved with 2 mM Mn(2+). K (m) and k (cat) values for diadenosine tetra-, penta- and hexaphosphates were 2.0, 2.4 and 1.1 microM and 11.4, 28.6 and 12.0 s(-1), respectively, while for GTP they were 20.3 microM and 1.8 s(-1), respectively. The K (m )for adenosine 5'-pentaphosphate was <1 microM. Expression analysis showed the DR2356 gene to be induced eight- to ninefold in stationary phase and in cells subjected to slow dehydration plus rehydration. Superoxide (but not peroxide) treatment and rapid dehydration caused a two-to threefold induction. The Mn-requirement and induction in stationary phase suggest that DR2356 may have a specific role in maintenance mode metabolism in stationary phase as Mn(2+) accumulates.
Fisher
David I
DI
School of Biological Sciences, Biosciences Building, University of Liverpool, P.O. Box 147, Liverpool, UK.
Cartwright
Jared L
JL
McLennan
Alexander G
AG
D017766:Acid Anhydride Hydrolases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D034301:Deinococcus / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D015226:Dinucleoside Phosphates / Q000737:chemistry / Q000378:metabolism; D004790:Enzyme Induction / Q000187:drug effects*; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000378:metabolism; D011122:Polyphosphates / Q000737:chemistry / Q000378:metabolism; D011755:Pyrophosphatases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D015290:Second Messenger Systems / Q000502:physiology; D016415:Sequence Alignment; D013379:Substrate Specificity; D013481:Superoxides / Q000494:pharmacology; D000096964:Nudix Hydrolases
D017766:Acid Anhydride Hydrolases; D001426:Bacterial Proteins; D015226:Dinucleoside Phosphates; D008345:Manganese; D009711:Nucleotides; D011122:Polyphosphates; D011755:Pyrophosphatases; D011994:Recombinant Proteins; D013481:Superoxides; D000096964:Nudix Hydrolases; C020733:diadenosine tetraphosphate; C075829:diadenosine polyphosphate hydrolase