Publications

• Identification of two human dimethylarginine dimethylaminohydrolases with distinct tissue distributions and homology with microbial arginine deiminases.

  Leiper J.M., Santa Maria J., Chubb A., MacAllister R.J., Charles I.G., Whitley G.S., Vallance P.

  Methylarginines inhibit nitric oxide synthases (NOS). Cellular concentrations of methylarginines are determined in part by the activity of dimethylarginine dimethylaminohydrolase (DDAH; EC 3.5.3. 18). We have cloned human DDAH and identified and expressed a second novel DDAH isoform (DDAH I and II ... >> More

  Methylarginines inhibit nitric oxide synthases (NOS). Cellular concentrations of methylarginines are determined in part by the activity of dimethylarginine dimethylaminohydrolase (DDAH; EC 3.5.3. 18). We have cloned human DDAH and identified and expressed a second novel DDAH isoform (DDAH I and II respectively). DDAH I predominates in tissues that express neuronal NOS. DDAH II predominates in tissues expressing endothelial NOS. These results strengthen the hypothesis that methylarginine concentration is actively regulated and identify molecular targets for the tissue and cell-specific regulation of methylarginine concentration. << Less

  Biochem. J. 343:209-214(1999) [PubMed] [EuropePMC]

• Mechanism of 4-HNE mediated inhibition of hDDAH-1: implications in NO regulation.

  Forbes S.P., Druhan L.J., Guzman J.E., Parinandi N., Zhang L., Green-Church K.B., Cardounel A.J.

  Nitric oxide synthase is inhibited by NG-methylated derivatives of arginine whose cellular levels are controlled by dimethylarginine dimethylamino-hydrolase (DDAH). DDAH-1 is a Zn(II)-containing enzyme that through hydrolysis of methylated l-arginines regulates the activity of NOS. Herein, we repo ... >> More

  Nitric oxide synthase is inhibited by NG-methylated derivatives of arginine whose cellular levels are controlled by dimethylarginine dimethylamino-hydrolase (DDAH). DDAH-1 is a Zn(II)-containing enzyme that through hydrolysis of methylated l-arginines regulates the activity of NOS. Herein, we report the kinetic properties of hDDAH-1 and its redox-dependent regulation. Kinetic studies using recombinant enzyme demonstrated Km values of 68.7 and 53.6 microM and Vmax values of 356 and 154 nmols/mg/min for ADMA and L-NMMA, respectively. This enzymatic activity was selective for free ADMA and L-NMMA and was incapable of hydrolyzing peptide incorporated methylarginines. Subsequent studies performed to determine the effects of reactive oxygen and reactive nitrogen species on DDAH activity demonstrated that low level oxidant exposure had little effect on enzyme activity and that concentrations approaching >or=100 microM were needed to confer significant inhibition of DDAH activity. However, exposure of DDAH to the lipid oxidation product, 4-HNE, dose-dependently inhibited DDAH activity with 15% inhibition observed at 10 microM, 50% inhibition at 50 microM, and complete inhibition at 500 microM. Mass spectrometry analysis demonstrated that the mechanism of inhibition resulted from the formation of Michael adducts on His 173, which lies within the active site catalytic triad of hDDAH-1. These studies were performed with pathophysiologicaly relevant concentrations of this lipid peroxidation product and suggest that DDAH activity can be impaired under conditions of increased oxidative stress. Because DDAH is the primary enzyme involved in methylarginine metabolism, the loss of activity of this enzyme would result in impaired NOS activity and reduced NO bioavailability. << Less

  Biochemistry 47:1819-1826(2008) [PubMed] [EuropePMC]

• Developing dual and specific inhibitors of dimethylarginine dimethylaminohydrolase-1 and nitric oxide synthase: toward a targeted polypharmacology to control nitric oxide.

  Wang Y., Monzingo A.F., Hu S., Schaller T.H., Robertus J.D., Fast W.

  Molecules that block nitric oxide's (NO) biosynthesis are of significant interest. For example, nitric oxide synthase (NOS) inhibitors have been suggested as antitumor therapeutics, as have inhibitors of dimethylarginine dimethylaminohydrolase (DDAH), an enzyme that catabolizes endogenous NOS inhi ... >> More

  Molecules that block nitric oxide's (NO) biosynthesis are of significant interest. For example, nitric oxide synthase (NOS) inhibitors have been suggested as antitumor therapeutics, as have inhibitors of dimethylarginine dimethylaminohydrolase (DDAH), an enzyme that catabolizes endogenous NOS inhibitors. Dual-targeted inhibitors hold promise as more effective reagents to block NO biosynthesis than single-targeted compounds. In this study, a small set of known NOS inhibitors are surveyed as inhibitors of recombinant human DDAH-1. From these, an alkylamidine scaffold is selected for homologation. Stepwise lengthening of one substituent converts an NOS-selective inhibitor into a dual-targeted NOS/DDAH-1 inhibitor and then into a DDAH-1 selective inhibitor, as seen in the inhibition constants of N5-(1-iminoethyl)-, N5-(1-iminopropyl)-, N5-(1-iminopentyl)- and N(5)-(1-iminohexyl)-l-ornithine for neuronal NOS (1.7, 3, 20, >1,900 microM, respectively) and DDAH-1 (990, 52, 7.5, 110 microM, respectively). A 1.9 A X-ray crystal structure of the N5-(1-iminopropyl)-L-ornithine:DDAH-1 complex indicates covalent bond formation between the inhibitor's amidino carbon and the active-site Cys274, and solution studies show reversible competitive inhibition, consistent with a reversible covalent mode of DDAH inhibition by alkylamidine inhibitors. These represent a versatile scaffold for the development of a targeted polypharmacological approach to control NO biosynthesis. << Less

  Biochemistry 48:8624-8635(2009) [PubMed] [EuropePMC]

• Purification and properties of a new enzyme, NG,NG-dimethylarginine dimethylaminohydrolase, from rat kidney.

  Ogawa T., Kimoto M., Sasaoka K.

  A new enzyme, NG, NG-dimethylarginine dimethylaminohydrolase which plays a role in the metabolism of NG,NG-dimethyl-L-arginine, has been purified to homogeneity from rat kidney. The enzyme consists of a single polypeptide and its molecular weight is about 33,000. The isoelectric point of the enzym ... >> More

  A new enzyme, NG, NG-dimethylarginine dimethylaminohydrolase which plays a role in the metabolism of NG,NG-dimethyl-L-arginine, has been purified to homogeneity from rat kidney. The enzyme consists of a single polypeptide and its molecular weight is about 33,000. The isoelectric point of the enzyme is at pH 5.2. The enzyme catalyzes the hydrolytic liberation of the dimethylamino moiety of NG,NG-dimethyl-L-arginine and forms L-citrulline and dimethylamine. It is highly specific for NG,NG-dimethyl-L-arginine and NG-monomethyl-L-arginine, and the Km values for these amino acids are 0.18 and 0.36 mM, respectively. The enzyme shows the maximum activity at pH 6.5 and requires no cofactor. The activity is strongly inhibited by SH-blocking reagents (e.g. p-chloromercuribenzoate and HgCl2) and divalent metal ions (e.g. Cd2+, Cu2+, and Zn2+). << Less

  J Biol Chem 264:10205-10209(1989) [PubMed] [EuropePMC]

• Characterization of dimethylargininase from bovine brain: evidence for a zinc binding site.

  Bogumil R., Knipp M., Fundel S.M., Vasak M.

  Dimethylargininase (EC 3.5.3.18) is involved in the regulation of the levels of the natural occurring free arginine derivatives L-Nomega,Nomega-dimethylarginine and L-Nomega-methylarginine, which are reversible inhibitors of nitric oxide synthase. A dimethylargininase has been isolated from bovine ... >> More

  Dimethylargininase (EC 3.5.3.18) is involved in the regulation of the levels of the natural occurring free arginine derivatives L-Nomega,Nomega-dimethylarginine and L-Nomega-methylarginine, which are reversible inhibitors of nitric oxide synthase. A dimethylargininase has been isolated from bovine brain tissue and was characterized by using immunological, kinetic, and spectroscopic techniques. Western blot analysis using polyclonal antibodies revealed that the enzyme is widely distributed in bovine with the highest relative concentrations found in brain and kidney tissue. A similar tissue distribution has also been reported for the other so far isolated dimethylargininase from rat kidney [Ogawa, T., Kimoto, M., and Sasaoka, K. (1989) J. Biol. Chem. 264, 10205-10209]. The bovine enzyme is a monomeric, globular protein (molecular mass approximately 31.2 kDa) containing one tightly bound Zn2+ ion, which can be removed by dialysis against 1,10-phenanthroline. The determination of kinetic constants for both the native (holo-protein) and the zinc-depleted (apo-protein) enzyme at 37 degrees ¿C established that the dimethylargininase is not a zinc hydrolase. The specific activity was 0.66 unit/mg for the holo-protein and 0.19 unit/mg for the apo-protein. The secondary structure determination of the native enzyme by circular dichroism revealed 41% alpha-helix and 32% beta-sheet and beta-turn structure. In the apo-enzyme, a small, but significant decrease in the alpha-helical content (5%) was observed, consistent with a marked decrease in enzymatic activity to 30%. Upon preincubation of both enzyme forms at 50 degrees C, only the holo-enzyme showed a residual enzymatic activity. In thermostability studies, a 7 degrees C lower apparent Tm value was observed for the apo-enzyme compared to the 66 degrees C for the holo-enzyme, suggesting that the zinc ion has a structure-stabilizing role. Besides the tightly bound zinc, additional Zn2+ ions inhibit the enzyme competitively with a Ki value of 2.0 microM. A possible interrelationship between dimethylargininase and nitric oxide synthase is discussed. << Less

  Biochemistry 37:4791-4798(1998) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.