Publications

• A rat liver lysosomal membrane flavin-adenine dinucleotide phosphohydrolase: purification and characterization.

  Shin H.J., Mego J.L.

  An enzyme hydrolyzing flavin-adenine dinucleotide (FAD) to flavin mononucleotide and AMP was identified and purified from rat liver lysosomal (Tritosomal) membranes. The purified enzyme showed a single band on silver-stained denaturing gels with an apparent Mr 70,000. Periodate-Schiff staining aft ... >> More

  An enzyme hydrolyzing flavin-adenine dinucleotide (FAD) to flavin mononucleotide and AMP was identified and purified from rat liver lysosomal (Tritosomal) membranes. The purified enzyme showed a single band on silver-stained denaturing gels with an apparent Mr 70,000. Periodate-Schiff staining after denaturing gel electrophoresis of whole membrane preparations revealed that this enzyme is one of the major glycoproteins in lysosomal membranes. FAD appeared to be the preferred substrate for the purified enzyme; equivalent concentrations of NAD or CoA were hydrolyzed at about one-half of the FAD rate. Negligible activity (less than or equal to 16%) was noted with ATP, TTP, ADP, AMP, FMN, pyrophosphate, or p-nitrophenylphosphate. The enzyme was inhibited by EDTA or dithiothreitol. It was stimulated by Zn, and was not affected by Ca or Mg ions, nor by p-chloromercuribenzoate. The pH optimum for FAD hydrolysis was 8.5-9 with an apparent Km of 0.125 mM. Antibodies prepared against the purified enzyme partially (50%) inhibited FAD phosphohydrolase activity in lysosomal membrane preparations but had no effect on the soluble lysosomal acid pyrophosphatase known to hydrolyze FAD. This enzyme could not be detected immunochemically in preparations of microsomes, Golgi, plasma membranes, mitochondrial membranes, or the soluble lysosomal fraction, suggesting that the enzyme is different from either soluble lysosomal acid pyrophosphatase or other FAD hydrolyzing activities in the liver cell. << Less

  Arch Biochem Biophys 267:95-103(1988) [PubMed] [EuropePMC]

• Molecular characterization of organelle-type Nudix hydrolases in Arabidopsis.

  Ogawa T., Yoshimura K., Miyake H., Ishikawa K., Ito D., Tanabe N., Shigeoka S.

  Nudix (for nucleoside diphosphates linked to some moiety X) hydrolases act to hydrolyze ribonucleoside and deoxyribonucleoside triphosphates, nucleotide sugars, coenzymes, or dinucleoside polyphosphates. Arabidopsis (Arabidopsis thaliana) contains 27 genes encoding Nudix hydrolase homologues (AtNU ... >> More

  Nudix (for nucleoside diphosphates linked to some moiety X) hydrolases act to hydrolyze ribonucleoside and deoxyribonucleoside triphosphates, nucleotide sugars, coenzymes, or dinucleoside polyphosphates. Arabidopsis (Arabidopsis thaliana) contains 27 genes encoding Nudix hydrolase homologues (AtNUDX1 to -27) with a predicted distribution in the cytosol, mitochondria, and chloroplasts. Previously, cytosolic Nudix hydrolases (AtNUDX1 to -11 and -25) were characterized. Here, we conducted a characterization of organelle-type AtNUDX proteins (AtNUDX12 to -24, -26, and -27). AtNUDX14 showed pyrophosphohydrolase activity toward both ADP-ribose and ADP-glucose, although its K(m) value was approximately 100-fold lower for ADP-ribose (13.0+/-0.7 microm) than for ADP-glucose (1,235+/-65 microm). AtNUDX15 hydrolyzed not only reduced coenzyme A (118.7+/-3.4 microm) but also a wide range of its derivatives. AtNUDX19 showed pyrophosphohydrolase activity toward both NADH (335.3+/-5.4 microm) and NADPH (36.9+/-3.5 microm). AtNUDX23 had flavin adenine dinucleotide pyrophosphohydrolase activity (9.1+/-0.9 microm). Both AtNUDX26 and AtNUDX27 hydrolyzed diadenosine polyphosphates (n=4-5). A confocal microscopic analysis using a green fluorescent protein fusion protein showed that AtNUDX15 is distributed in mitochondria and AtNUDX14 -19, -23, -26, and -27 are distributed in chloroplasts. These AtNUDX mRNAs were detected ubiquitously in various Arabidopsis tissues. The T-DNA insertion mutants of AtNUDX13, -14, -15, -19, -20, -21, -25, -26, and -27 did not exhibit any phenotypical differences under normal growth conditions. These results suggest that Nudix hydrolases in Arabidopsis control a variety of metabolites and are pertinent to a wide range of physiological processes. << Less

  Plant Physiol. 148:1412-1424(2008) [PubMed] [EuropePMC]