Makoto Shimoyama

Effect of polyamines on phosphorylation of non-histone chromatin proteins from hog liver

Abstract The effects of polyamines on the in vitro phosphorylation of non-histone chromatin proteins from hog liver has been found to be dose dependent. Maximal increase occurred at 0.2 mM spermine and 2 mM spermidine, respectively. These results suggest that spermine and spermidine may have a regulating function for phosphorylation of non-histone chromatin proteins in hog liver.

Nicotinamide inhibition of 3′,5′- cyclic AMP phosphodiesterase in vitro

Abstract Nicotinamide was found to be potent inhibitor of 3′,5′- cyclic AMP phosphodiesterase, but nicotinamide adenine dinucleotide was found impotent. Therefore, it may be presumed that the induction of certain enzymes following the intraperitoneal injection of nicotinamide into rats would due to the elevation of the steady state level of 3′,5′- cyclic AMP through the inhibition of phosphodiesterase by nicotinamide.

Identification of the 3′, 5′- cyclic AMP phosphodiesterase inhibitor in potato: Feed-back control by inorganic phosphate

Abstract We attempted to identify the low molecular sized 3′, 5′- cyclic AMP phosphodiesterase inhibitor in potato. Finding that the partially purified inhibitor yielded an elution pattern and a phosphodiesterase inhibition identical with those of inorganic phosphate in Sephadex G-25 column chromatography, and finding that the Rf value for this inhibitor agreed with that for inorganic phosphate on paper chromatographic analysis, we are able to identify this inhibitor as inorganic phosphate, a known end-product of cyclic AMP catabolism.

Evidence for and some properties of a 3′, 5′- cyclic AMP phosphodiesterase inhibitor in potato

Summary Dialysis of potato 105,000 × g supernatant resulted in a dramatic increase in 3′, 5′- cyclic AMP phosphodiesterase activity, due to the removal of some unknown inhibitor of small molecular size, through this process. The inhibitor obtained by gel filtration was resistant to boiling at neutral, acidic, or alkaline pH and also to charcoal treatment. Kinetic analysis showed that inhibition was dependent on inhibitor concentration in the reaction mixture and that it was noncompetitive.

NAD glycohydrolase from the small intestine of the rat: Purification, properties and possible role of the enzyme

Abstract 1. 1. In the investigation of NAD glycohydrolase (EC 3.2.2.5) in the small intestine of the rat, the properties of this enzyme solubilized by lipase (glycerol-ester hydrolase, EC 3.1.1.3) and purified 250-fold were found to be similar in essentials to its properties found in other tissues. The Km values for NAD and NADP were 2.7·10−5 and 3.8·10−6 M, respectively. 2. 2. An over 70% inhibition of NAD hydrolysis was observed in the presence of less than 1·10−6 M NADP when NAD was used at a concentration of 6.25·10−6 M as substrate. In contrast to the NADP effect on the NAD cleavage, a slight effect of NAD on the hydrolysis of the NADP was observed. 3. 3. The permeability of pyridine nucleotide in the small intestine was studied. No penetration of [14C]NAD prior to degradation to [14C]nicotinamide was observed. This result would indicate that NAD glycohydrolase may contribute to the absorption of the pyridine moiety of NAD, which would be found significant in the diet.

Nicotinamide adenine dinucleotide in developing chick embryo

Abstract The changes in NAD levels during the development of the chick embryo were investigated. Almost all NAD in the fertilized egg before incubation was localized in the white albumin, and NAD decreased rapidly during incubation. The hepatic NAD levels varied significantly during the development of the chick embryo. When several NAD precursors such as nicotinamide, nicotinic acid and tryptophan were injected into fertilized eggs before incubation, an increase of NAD in chick embryo was found in the first and middle periods of embryonic development, but no increase was observed in the latter stage.

Nicotinamide-adenine dinucleotide pyrophosphatase of Cambaroides japonica

Abstract 1. 1.The cleavage of NAD in crayfish hepatopancreas is catalyzed chiefly by a pyrophosphatase rather than by NAD glycohydrolase (EC 3.2.2.5). This fact was confirmed by the loss in the coenzyme function of NAD for yeast alcohol dehydrogenase without a significant concomitant loss in reactivity towards cyanide and the identification of the reaction products as NMN and AMP by means of paper chromatography and Dowex 1-X2 column chromatography. 2. 2.NAD pyrophosphatase is localized chiefly in mitochondria and microsomes. The enzyme was partially purified by (NH 4 ) 2 SO 4 fractionation. Using this preparation, the K m value for NAD was determined as 1.1·10 −3 M. The activity with reduced NAD was about 3-fold higher than with NAD. ATP and NADP are not cleaved by the crayfish enzyme. 3. 3.NAD pyrophosphatase is inactivated by heating at 60° for 1 min and no stimulation was observed by heating at 40° for 1 min. The pH optimum is 8.4. 4. 4.Enzyme activity is inhibited by mononucleotides such as AMP, GMP and UMP, and AMP inhibition is competitive with substrate. Nicotinamide did not inhibit the enzyme activity at 1·10 −3 M. 5. 5.The possible metabolic significance of AMP inhibition of NAD hydrolyzing enzymes in mammalian tissues is also discussed.

Effect of polyanions and polycations on adenosine 3′,5′- monophosphate-binding and protein kinase activity

Abstract Histone, protamine, poly-L-arginine, and poly-L-lysine enhance the binding of adenosine 3′,5′-monophosphate (cyclic AMP) to rat liver cyclic AMP-dependent protein kinase as determined by Millipore filtration assay. Poly-L-glutamic acid and poly-L-aspartic acid suppress cyclic AMP-binding stimulated by histone. Poly-L-glutamic acid and poly-L-aspartic acid are effective against protein kinase and result in decrease in initial reaction velocity when histone is used as a protein substrate. Incubation of cyclic AMP-dependent protein kinase with 6 μg poly-L-glutamic acid produces half-maximal inhibition of cyclic AMP-dependent protein kinase when 30 μg histone is used as substrate.

Subcellular localization of nicotinamide deamidase in rat liver

Abstract Trough the isolation of subcellular structural components in a state free from cross-contamination, nicotinamide deamidase in rat liver has been found to be localized solely in the microsomes. Furthermore, no nicotinamide deamidase activity was detected in isolated plasma membrane. Isolated mitochondria contained negligible amounts of nicotinamide deamidase activity, which suggests that the previously reported substantial recovery of the deamidase in the mitochondrial fraction2 must have been due to contaminating microsomes.

Target Proteins for Arginine-Specific ADP-Ribosyltransferase in Chickens

Eukaryotic arginine-specific ADP-ribosyltransferases are present in various types of tissues and cells. Although some of the related enzymes have been purified and characterized (Moss et al., 1980, Tanigawa et al., 1984, Peterson et al., 1990, Larew et al., 1991) the physiological role of the enzymes remains obscure, presumably because much less is known of the endogenous acceptor protein. Identification and characterization of target proteins for endogenous arginine-specific ADP-ribosylation provide clues to the physiological function of ADP-ribosylation.