3,4-Dihydrocoumarin hydrolase with haloperoxidase activity from Acinetobacter calcoaceticus F46. (1/16)

A novel lactonohydrolase, an enzyme that catalyzes the hydrolysis of 3,4-dihydrocoumarin, was purified 375-fold to apparent homogeneity, with a 22.7% overall recovery, from Acinetobacter calcoaceticus F46, which was isolated as a fluorene-assimilating micro-organism. The molecular mass of the native enzyme, as estimated by high-performance gel-permeation chromatography, is 56 kDa, and the subunit molecular mass is 30 kDa. The enzyme specifically hydrolyzes 3,4-dihydrocoumarin, and the Km and Vmax for 3,4-dihydrocoumarin are 0.806 mM and 4760 U.mg-1, respectively. The N-terminal and internal amino acid sequences of the enzyme show high similarity to those of bacterial non-heme haloperoxidases. The enzyme exhibits brominating activity with monochlorodimedon in the presence of H2O2 and 3, 4-dihydrocoumarin or an organic acid, such as acetate and n-butyrate.  (+info)

Al-induced efflux of organic acid anions is poorly associated with internal organic acid metabolism in triticale roots. (2/16)

The secretion of organic acid anions from roots has been identified as a mechanism of resistance to Al. However, the process leading to the secretion of organic acid anions is poorly understood. The effect of Al on organic acid metabolism was investigated in two lines of triticale (xTriticosecale Wittmark) differing in Al-induced secretion of malate and citrate and in Al resistance. The site of Al-induced secretion of citrate and malate from a resistant line was localized to the root apices (terminal 5 mm). The levels of citrate (root apices and mature root segments) and malate (mature segments only) in roots increased during exposure to Al, but similar changes were observed in both triticale genotypes. The in vitro activities of four enzymes involved in malate and citrate metabolism (citrate synthase, phosphoenolpyruvate carboxylase, malate dehydrogenase, and NADP-isocitrate dehydrogenase) were similar for sensitive and resistant lines in both root apices and mature root segments. The response of these enzymes to pH did not differ between tolerant and sensitive lines or in the presence and absence of Al. Moreover, cytoplasmic and vacuolar pH were not affected by exposure to Al in either line. Together, these results indicate that the Al-dependent efflux of organic acid anions from the roots of triticale is not regulated by their internal levels in the roots or by the capacity of the root cells to synthesize malate and citrate.  (+info)

Propionyl-coenzyme A synthetases of Ralstonia solanacearum and Salmonella choleraesuis display atypical kinetics. (3/16)

Propionyl-coenzyme A synthetases (PrpE) of Salmonella choleraesuis and Ralstonia solanacearum sharing 62% identity in amino acid sequence to each other were cloned, expressed in Escherichia coli and purified. Both enzymes catalyzed acetyl-, propionyl-, butyryl- and acrylyl-coenzyme A formation with the highest k(cat)/K(m) values for propionate. They displayed sigmoidal homotrophic autoactivation kinetics for propionate but not for the other acyl substrates tested. Besides, substrate inhibition kinetics was observed for co-substrates, i.e. ATP and CoA. Based on the kinetic data reported herein, the reaction mechanisms of the enzyme are discussed.  (+info)

The inhibition of the human cholesterol 7alpha-hydroxylase gene (CYP7A1) promoter by fibrates in cultured cells is mediated via the liver x receptor alpha and peroxisome proliferator-activated receptor alpha heterodimer. (4/16)

In previous work, we showed that the binding of the liver x receptor alpha:peroxisome proliferator-activated receptor alpha (LXRalpha:PPARalpha) heterodimer to the murine Cyp7a1 gene promoter antagonizes the stimulatory effect of their respective ligands. In this study, we determined if LXRalpha:PPARalpha can also regulate human CYP7A1 gene promoter activity. Co-expression of LXRalpha and PPARalpha in McArdle RH7777 hepatoma cells decreased the activity of the human CYP7A1 gene promoter in response to fibrates and 25-hydroxycholesterol. In vitro, the human CYP7A1 Site I bound LXRalpha:PPARalpha, although with substantially less affinity compared with the murine Cyp7a1 Site I. The binding of LXRalpha:PPARalpha to human CYP7A1 Site I was increased in the presence of either LXRalpha or PPARalpha ligands. In HepG2 hepatoblastoma cells, fibrates and 25-hydroxycholesterol inhibited the expression of the endogenous CYP7A1 gene as well as the human CYP7A1 gene promoter when co-transfected with plasmids encoding LXRalpha and PPARalpha. However, a derivative of the human CYP7A1 gene promoter that contains a mutant form of Site I that does not bind LXRalpha:PPARalpha was not inhibited by WY 14,643 or 25-hydroxycholesterol in both McArdle RH7777 and HepG2 cells. The ligand-dependent recruitment of LXRalpha:PPARalpha heterodimer onto the human CYP7A1 Site I can explain the inhibition of the human CYP7A1 gene promoter in response to fibrates and 25-hydroxycholesterol.  (+info)

Corn hybrid affects in vitro and in vivo measures of nutrient digestibility in dogs. (5/16)

Corn is a commonly used ingredient in dry pet foods because there is a stable supply and it is a relatively inexpensive source of nutrients. Corn hybrids are available that are higher in CP and amylose and lower in phytate concentration than conventional hybrids. Approximately 500 mg of high-protein (HP), high-protein, low-phytate (HPLP), and high-amylose (HA) corn were compared with conventional (CONV) corn and amylomaize starch (AM) in triplicate and exposed to pepsin/hydrochloric acid and pancreatin to simulate hydrolytic digestion. Substrate remaining after this was used to determine in vitro colonic fermentation. Organic matter disappearances as a result of hydrolytic digestion were >80% for CONV, HP, and HPLP, whereas HA (60.7%) and AM (43.7%) were lower (P < 0.05). Total digestion (TD) values after hydrolytic digestion and 8 h of fermentation using canine fecal inoculum were greater (P < 0.05) for CONV, HP, and HPLP vs. HA and AM. The residue left after hydrolytic digestion of all substrates was poorly fermented. Five ileal-cannulated dogs were fed each corn hybrid at approximately 31% of the diet in a 5 x5 Latin square design. Dogs fed diets containing HP corn had higher (P < 0.05) ileal OM digestibility (70.3%) and tended (P < 0.10) to have higher DM digestibility (64.6%). Ileal starch digestibilities were lower (P < 0.05) for dogs fed HA (64.0%) and AM (63.0%). Ileal digestibilities of essential (71.2%), nonessential (67.4%), and total (69.0%) AA tended to be higher (P < 0.10) for HP diets compared with CONV (66.4, 62.4, and 64.0%, respectively). Total-tract DM, OM, CP, and GE digestibilities (77, 82, 77, and 84%, on average, respectively) were higher (P < 0.05) for dogs fed CONV, HP, and HPLP than for those fed AM (66.9, 71.6, 72.6, and 76.5%) and HA (60.6, 65.7, 69.7, and 71.5%). Total-tract fat digestibilities were lower (P < 0.05) for dogs fed HA diets (86.6%) than for all other treatments (91.0%, on average). Total-tract starch digestibilities were higher (P < 0.05) for dogs fed CONV, HP, and HPLP (98%, on average) compared with HA (72.8%) and AM (76.5%). No differences were detected among treatments in fecal bifidobacteria, lactobacilli, or Clostridium perfringens concentrations. The experiments demonstrated that HP and HPLP corn had hydrolytic digestion and fermentation characteristics similar to those of CONV corn, whereas HA resulted in similar responses to AM, a well-established resistant starch ingredient.  (+info)

Enantioselective fluorescent recognition of chiral acids by cyclohexane-1,2-diamine-based bisbinaphthyl molecules. (6/16)

The cyclohexane-1,2-diamine-based bisbinaphthyl macrocycles (S)-/(R)-5 and their cyclic and acyclic analogues are synthesized. The interactions of these compounds with various chiral acids are studied. Compounds (S)-/(R)-5 exhibit highly enantioselective fluorescent responses and high fluorescent sensitivity toward alpha-hydroxycarboxylic acids and N-protected amino acids. Among these interactions, (S)-mandelic acid (10(-3) M) led to over 20-fold fluorescence enhancement of (S)-5 (1.0 x 10(-5) M in benzene/0.05% DME) at the monomer emission, and (S)-hexahydromandelic acid (10(-3) M) led to over 80-fold fluorescence enhancement. These results demonstrate that (S)-5 is useful as an enantioselective fluorescent sensor for the recognition of the chiral acids. On the basis of the study of the structures of (S)-5 and the previously reported 1,2-diphenylethylenediamine-based bisbinaphthyl macrocycle (S)-4, the large fluorescence enhancement of (S)-5 with a chirality-matched alpha-hydroxycarboxylic acid is attributed to the formation of a structurally rigidified host-guest complex and the further interaction of this complex with the acid to suppress the photoinduced electron-transfer fluorescent quenching caused by the nitrogens in (S)-5.  (+info)

Bioactive constituents from Chinese natural medicines. XXVIII. Chemical structures of acyclic alcohol glycosides from the roots of Rhodiola crenulata. (7/16)

Five new glycosides, creosides I, II, III, IV, and V, were isolated from the methanolic extract of the roots of Rhodiola crenulata, together with 21 known compounds. The chemical structures of new constituents were elucidated on the basis of chemical and physicochemical evidence.  (+info)

Mutation of the active site carboxy-lysine (K70) of OXA-1 beta-lactamase results in a deacylation-deficient enzyme. (8/16)

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