Propionaldehyde Dehydrogenase Research Articles

Purification, characterization and subunits identification of the diol dehydratase of Lactobacillus collinoides.

The three genes pduCDE encoding the diol dehydratase of Lactobacillus collinoides, have been cloned for overexpression in the pQE30 vector. Although the three subunits of the protein were highly induced, no activity was detected in cell extracts. The enzyme was therefore purified to near homogeneity by ammonium sulfate precipitation and gel filtration chromatography. In fractions showing diol dehydratase activity, three main bands were present after SDS/PAGE with molecular masses of 63, 28 and 22 kDa, respectively. They were identified by mass spectrometry to correspond to the large, medium and small subunits of the dehydratase encoded by the pduC, pduD and pduE genes, respectively. The molecular mass of the native complex was estimated to 207 kDa in accordance with the calculated molecular masses deduced from the pduC, D, E genes (61, 24.7 and 19,1 kDa, respectively) and a alpha2beta2gamma2 composition. The Km for the three main substrates were 1.6 mm for 1,2-propanediol, 5.5 mm for 1,2-ethanediol and 8.3 mm for glycerol. The enzyme required the adenosylcobalamin coenzyme for catalytic activity and the Km for the cofactor was 8 micro m. Inactivation of the enzyme was observed by both glycerol and cyanocobalamin. The optimal reaction conditions of the enzyme were pH 8.75 and 37 degrees C. Activity was inhibited by sodium and calcium ions and to a lesser extent by magnesium. A fourth band at 59 kDa copurified with the diol dehydratase and was identified as the propionaldehyde dehydrogenase enzyme, another protein involved in the 1,2-propanediol metabolism pathway.

Effects of β-naphthoflavone on the cytochrome P450 system, and phase II enzymes in gilthead seabream ( Sparus aurata)

The effect of β-naphthoflavone (β-NF) on several catalytic activities of cytochrome P450 (CYP) and phase II enzymes putatively controlled by [Ah]-receptor activation in the liver, heart and kidney of gilthead seabream, was investigated. In the liver, β-NF treatment [intraperitoneal injection (i.p.) 50 mg/kg] resulted in an increase of CYP content, immunoreactive CYP 1A and methoxyresorufin- O-demethylase (MEROD), pentoxyresorufin O-depentylase (PROD) and ethoxyresorufin- O-deethylase (EROD) activities. However, β-NF had no effect on any of the hepatic phase II enzymes examined (benzaldehyde dehydrogenase, propionaldehyde dehydrogenase, glutathione S-transferase, UDP-glucuronyl-transferase, DT-diaphorase). Single i.p. injection of 10 mg/kg β-NF showed a maximal induction of CYP 1A-like protein and EROD activity after 3–7 days. CYP 1A and EROD returned to control levels 18-days post-treatment. β-NF injection also caused a rapid increase of a single band size of mRNA recognized by a CYP 1A1 cDNA fragment from sea bass ( Dicentrarchus labrax). Expression of mRNA preceded the increase of EROD activity and declined rapidly by 96 h. Dose–response experiments demonstrated that EROD was significantly enhanced in liver by a single injection of 0.3 mg/kg β-NF and was the most sensitive measurement for CYP 1A-like induction. β-NF treatments also increased the expression of CYP 1A-like protein, mRNA and EROD, but not MEROD and PROD activities in heart and kidney.

Biochemical and histochemical responses to environmental contaminants in clam, Tapes philippinarum, transplanted to different polluted areas of Venice Lagoon, Italy

In this study the clam, Tapes philippinarum, a filter-feeding bivalve living in soft bottoms, was used to investigate the water/sediment pollution in Venice Lagoon (Venice, Italy), a heavily urbanised and industrialised area. To this end, clams collected from indigenous populations in a clean farming area (Val Dogà) were transplanted into a relatively low contaminated site (Palude della Rosa) and in a heavily polluted area (Porto Marghera), for 5 weeks. A range of possible induced stress indices were measured, on the basis of potential toxicity mechanisms, including specific enzymatic activities [Bap hydroxylase and NADPH cytochrome c reductase, superoxide dismutase, catalase, glutathione peroxidase and propionaldehyde dehydrogenase (PDH)] and general cellular responses (lysosomal latency). Moreover, chemical analyses (polychlorinated biphenyls, polycyclic aromatic hydrocarbon, DDTs) were carried out on sediments and animal tissues at the beginning and end of the transplantation experiments. The chemical results show an active bio-accumulation activity from sediment in clams translocated to the most polluted area. Correspondingly, biological data indicate marked effects on PDH activity and latency in T. philippinarum from the same site. In contrast, no similar pattern has been observed in the other considered indices, possibly due to antagonistic effects of the complex contaminant mixture present in the environment.

Effect of Clofibrate, a Peroxisome Proliferator, in Sea Bass (Dicentrarchus labrax), a Marine Fish

The effects of 35 or 70 mg/kg clofibrate on some peroxisomal, mitochondrial, and microsomal enzymes (markers of peroxisome proliferation) in liver, gill, and kidney of sea bass following 2 weeks i.p. treatment have been studied. Induction of lauric acid hydroxylase, UDP-glucuronyl transferase, palmitoyl-CoA oxidase, carnitine-palmitoyl-transferase,p-nitrophenylacetate hydrolase, and benzaldehyde and propionaldehyde dehydrogenase activities was not observed in any case. The clofibrate administration at the dose of 70 mg/kg induced in the liver the glutathioneS-tranferase and reduced epoxide hydrolase activities. These results demonstrate that a marine fish species, such as sea bass, is rafractary to peroxisome proliferation.

Assessment of the impact of chemical pollutants on mussel, Mytilus galloprovincialis, from the Venice Lagoon, Italy

The purpose of this study was to identify possible relationships between biochemical—Catalase, Superoxide dismutase (SOD) and Propionaldehyde dehydrogenase (PDH)—and histochemical—Neutral Red Retention time (NRR)—parameters of mussels ( Mytilus galloprovincialis) and chemical (Hg, Pb, Cd, Zn, Cr, DDTs and PCBs) contaminants from the most polluted area of the Venice Lagoon which lies around Porto Marghera (industrial pollution) and the city of Venice (domestic pollution). Sampling of mussels was undertaken at the beginning of March 1995 and repeated in March 1996, just before spawning, when tissue concentrations of chemical pollutants reach their annual maxima. The results of the organic contaminant analysis showed generally good agreement between sites for the two sampling periods, with values broadly 10% lower during 1996 as compared to 1995. Metal contaminant concentrations in tissues revealed greater variability than organic compounds. Distribution pattern of biological parameters were more complex and variable. The apparent lack of significant positive correlations between antioxidant enzyme activities and pollutant body burdens as well as the low values of NRR are consistent with the present debate on the use of these biological indices as markers of contaminant-mediated stress for molluscs.

Genetic characterization of the pdu operon: use of 1,2-propanediol in Salmonella typhimurium.

Salmonella typhimurium is able to catabolize 1,2-propanediol for use as the sole carbon and energy source; the first enzyme of this pathway requires the cofactor adenosyl cobalamin (Ado-B12). Surprisingly, Salmonella can use propanediol as the sole carbon source only in the presence of oxygen but can synthesize Ado-B12 only anaerobically. To understand this situation, we have studied the pdu operon, which encodes proteins for propanediol degradation. A set of pdu mutants defective in aerobic degradation of propanediol (with exogenous vitamin B12) defines four distinct complementation groups. Mutations in two of these groups (pduC and pduD) eliminate propanediol dehydratase activity. Based on mutant phenotypes, a third complementation group (pduG) appears to encode a cobalamin adenosyl transferase activity. No function has been assigned to the pduJ complementation group. Propionaldehyde dehydrogenase activity is eliminated by mutations in any of the four identified complementation groups, suggesting that this activity may require a complex of proteins encoded by the operon. None of the mutations analyzed affects either of the first two genes of the operon (pduA and pduB), which were identified by DNA sequence analysis. Available data suggest that the pdu operon includes enough DNA for about 15 genes and that the four genetically identified genes are the only ones required for aerobic use of propanediol.

Responses of hepatic biotransformation and antioxidant enzymes to CYP1A-inducers (3-methylcholanthrene, β-naphthoflavone) in sea bass ( Dicentrarchus labrax), dab ( Limanda limanda) and rainbow trout ( Oncorhynchus mykiss)

The time-course responses of hepatic biotransformation and antioxidant enzymes to cytochrome P4501A-inducers in fish were examined using marine and freshwater species, viz. respectively, sea bass ( Dicentrarchus labrax) and dab ( Limanda limanda) exposed up to 5 to 11 days to 3-methylcholanthrene (3MC; intraperitoneal (i.p.) 20 mg kg −1), and rainbow trout ( Oncorhynchus mykiss) exposed up to 15 days to β-naphthoflavone (BNF; i.p. 5 mg kg −1). The enzymes included those which are part of the mammalian [ Ah]-gene locus, viz. cytochrome P4501A (CYP1A), DT-diaphorase (DTD; EC 1.6.99.2), aldehyde dehydrogenase (ADH; EC 1.2.1.3), glutathione S-transferase (GST; EC 2.5.1.18) and UDP-glucuronyltransferase (UDPGT). DTD was assayed as dicoumarol-inhibitable menadione reductase, dicoumarol-inhibitable dichlorophenolindophenol (DCPIP) reductase and resorufin reductase activities; and ADH as benzaldehyde dehydrogenase (BADH) and propionaldehyde dehydrogenase (PADH) activities. The responses varied considerably with enzyme, species and treatment. CYP1A-dependent 7-ethoxyresorufin O-deethylase (EROD) activity was markedly inducible, more so in BNF-treated O. mykiss (maximal 284-fold increase after 2 days) than the 3MC-treated species (maximal 24–30-fold). Modest increases (up to 2-fold) and/or inhibition were seen in UDPGT and GST activities. DTD activities increased about 2-fold after 8 to 15 days in BNF-treated O. mykiss, but were not elevated in the other species. Changes in dicoumarol-inhibitable DCPIP and menadione reductase activities were more similar than resorufin reductase activity. No marked increases were evident in BADH, PADH, superoxide dismutase (SOD; EC 1.15.1.1), catalase (EC 1.11.1.6) and glutathione peroxidase (GPX; EC 1.11.1.9) activities in any species. Overall, some co-induction of CYP1A and DTD, over different time-courses, was indicated for BNF-treated O. mykiss.

Marginal role of impaired aldehyde metabolism in m‐xylene vapour‐induced biochemical effects in the rat

AbstractRats were exposed to 300 ppm m‐xylene vapour for 1 or 2 weeks, 6 h daily, 5 days a week. One group was simultaneously given disulfiram in the drinking water (53 μmol kg−1, daily). The disulfiram treatment inhibited propionaldehyde dehydrogenase; such inhibition did not occur when m‐methylbenzaldehyde (i.e., the aldehyde of m‐xylene metabolism) was used as the substrate. Gastric gavage with higher drug doses (0.5 g kg−1, administered one or three times) confirmed this finding. Intermittent inhalation exposure to m‐xylene for 2 weeks lowered the non‐protein sulphhydryl group content in liver and increased the activities of microsomal NADPH‐cytochrome c reductase, 7‐ethoxycoumarin O‐deethylase and UDP‐glucuronosyltransferase in liver. Concomitantly, the oxidation of m‐xylene in vitro was also enhanced. Combined exposure to the solvent and the drug somewhat inhibited the increases in the hepatic drug metabolism and in the rate of microsomal xylene oxidation in vitro which m‐xylene had induced, whereas the renal drug metabolism remained almost unaffected. Despite the changes in the monooxygenase, the solvent burden in the perirenal fat remained comparable. The significance of impaired aldehyde metabolism in m‐xylene toxicity could not be confirmed in male Wistar rats at the above exposure levels.