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Tetsuko Takabe - One of the best experts on this subject based on the ideXlab platform.
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An isozyme of betaine aldehyde Dehydrogenase in barley.
Plant & cell physiology, 2001Co-Authors: Toshihide Nakamura, Mika Nomura, Hitoshi Mori, Andre T. Jagendorf, Akihiro Ueda, Tetsuko TakabeAbstract:Betaine aldehyde Dehydrogenase (BADH) is an important enzyme for Gly betaine synthesis. We isolated two types of BADH cDNAs (BBD1 and BBD2) from barley. As BBD1 contained the signal sequence (SKL) targeting to microbodies, BBD2 was more similar to previously reported genes coding for BADH in dicotyledons (chloroplast type) than those in monocotyledons (microbody type). The two barley BADH genes showed different expression patterns. The BBD1 transcript was more abundant in roots than leaves and was induced to higher levels by salt, drought and abscisic acid (ABA) treatment. BBD2 transcript was more abundant in leaves and induced by salt, drought, PEG and ABA treatment. To understand the processing of these BADH proteins, we partially purified both enzymes and determined their N-terminal sequences. Based on comparisons of the N-terminal sequences to their deduced amino acid sequence, neither BBD1 nor BBD2 is processed at the N-terminus. These results suggest that BBD2 codes for a new type of BADH, which is not localized in either chloroplasts or mitochondria.
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molecular cloning and functional characterization of two kinds of betaine aldehyde Dehydrogenase in betaine accumulating mangrove avicennia marina forsk vierh
Plant Molecular Biology, 2001Co-Authors: Takashi Hibino, Tetsuko Takabe, Nobuyuki Matsuda, Yu-ling Meng, Hiroshi Ishikawa, Yoshinobu Kawamitsu, N Uehara, Yoshito Tanaka, Shigeyuki Baba, Keishiro WadaAbstract:Glycinebetaine is an important osmoprotectant in bacteria, plants, and animals, but only little information is available on the synthesis of glycinebetaine in tree plants. Among four mangrove species, glycinebetaine could be detected only in Avicennia marina. Pinitol was the main osmoprotectant in the other three species. The level of glycinebetaine in A. marina increased under high salinity. Betaine-Aldehyde Dehydrogenase (BADH) was detected in all four species, but choline monooxygenase could not be detected. A cDNA library was constructed from the leaves of A. marina. Two kinds of BADH cDNAs were isolated, one homologous to the spinach chloroplast BADH, and the other with unique residues SKL at the end of C-terminus. The BADH transcription levels of the former were higher than those of the latter. The levels of the former BADH increased at high salinity whereas those of the latter were independent of salinity. BADHs were expressed in Escherichia coli and purified. Two kinds of A. marina BADHs exhibited similar kinetic and stability properties, but were significantly different from those of spinach BADH. A. marina BADHs efficiently catalyzed the oxidation of betainealdehyde, but not the oxidation of omega-aminoaldehydes and were more stable at high temperature than the spinach BADH.
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Overproduction of spinach betaine aldehyde Dehydrogenase in Escherichia coli
European journal of biochemistry, 2000Co-Authors: Aran Incharoensakdi, Tetsuko Takabe, Nobuyuki Matsuda, Takashi Hibino, Yu-ling Meng, Hiroshi Ishikawa, Akira Hara, Tohru Funaguma, Teruhiro TakabeAbstract:Betaine aldehyde Dehydrogenase (BADH) catalyzes the last step in the synthesis of the osmoprotectant glycine betaine from choline. Although betaine aldehyde has been thought to be a specific substrate for BADH, recent studies have shown that human and sugar beet BADHs also catalyze the oxidation of ω-aminoaldehydes. To characterize the kinetic and stability properties of spinach BADH, five kinds of expression vectors encoding full length, mature, E103Q, E103K, and chimera BADHs were constructed. These enzymes together with Escherichia coli BADH were expressed in E. coli and purified. The affinities for betaine aldehyde were similar in the spinach and E. coli BADHs, whereas those for ω-aminoaldehydes were higher in spinach BADH than in E. coli BADH. A chimera BADH in which part of the Rossmann type fold in the spinach BADH was replaced with that of E. coli BADH, showed properties which resembled spinach BADH more than E. coli BADH. The spinach E103K mutant was almost inactive, whereas the E103Q mutant showed a similar activity for the oxidation of betaine aldehyde to that of wild type BADH, but a lower affinity for ω-aminoaldehydes. All spinach BADHs were dimers whereas E. coli BADH was a tetramer. E. coli BADH was more stable at high temperature than spinach BADHs. The E103Q mutant was most labile to high temperature. These properties are discussed in relation to the structure of spinach BADH.
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Compatibility of glycinebetaine in rice plants: evaluation using transgenic rice plants with a gene for peroxisomal betaine aldehyde Dehydrogenase from barley
Plant Cell & Environment, 2000Co-Authors: Sachie Kishitani, T. Takanami, M. Suzuki, M. Oikawa, Shuji Yokoi, Manabu Ishitani, A. M. Alvarez‐nakase, Tetsuko TakabeAbstract:Glycinebetaine is synthesized in plants by the two-step oxidation of choline, with betaine aldehyde as the intermediate. The reactions are catalyzed by choline mono-oxygenase and betaine aldehyde Dehydrogenase. Rice plants, which do not accumulate glycinebetaine, possess a gene encoding betaine aldehyde Dehydrogenase, whose activity is detectable at low levels. To evaluate the compatibility in rice of glycinebetaine on growth and tolerance to salt, cold and heat, we produced transgenic rice plants by introduction of a cDNA for betaine aldehyde Dehydrogenase of barley, which is localized in peroxisomes unlike the chloroplast-specific localization of betaine aldehyde Dehydrogenase in spinach and sugar beet. The transgenic rice plants converted high levels of exogenously applied betaine aldehyde (up to 10 mol m 3 ) to glycinebetaine more efficiently than did wild-type plants. The elevated level of glycinebetaine in transgenic plants conferred significant tolerance to salt, cold and heat stress. However, very high levels of glycinebetaine, resulting from conversion of applied betaine aldehyde to glycinebetaine or from exogenous application, inhibited increases in length of rice plants but not increases in dry weight. Our results suggested that the benefits of accumulation of glycinebetaine by rice plants might be considerable under high light conditions.
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isolation of a barley gene encoding betaine aldehyde Dehydrogenase that is not localized in microbody
1998Co-Authors: Toshihide Nakamura, Mika Nomura, Maurice S B Kul, Tetsuko TakabeAbstract:In some higher plants, glycinebetaine is synthesized and accumulated in cells in response to salt stress. Glycinebetame is a compatible solute and acts as an excellent osmoprotectant. High levels of glycinebetame arc present in leaves of diverse families of dicotyledons (1) and of some monocotyledons (2). The pathway of glycinebetaine biosynthesis in higher plants is as follows : Choline Betaine aldehyde → Glycine betaine. The enzyme which catalyzes the first step is choline monooxygenase (CMO), being recently purified and characterized (3,4). The final step is catalyzed by betaine aldehyde Dehydrogenase (BADH), which has been well characterized (5,6,7). Existence of B'ADH isozyme was suggested in spinach and sorghum (8,9), and we reported that all monocotyledonous BADHs have a C-terminal tripeptide SKL that is known to be a signal for targeting preproteins to microbodies (10).
Elisa M. Valenzuela-soto - One of the best experts on this subject based on the ideXlab platform.
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Effect of the drug cyclophosphamide on the activity of porcine kidney betaine aldehyde Dehydrogenase
Molecular and Cellular Biochemistry, 2021Co-Authors: Ramses Cruz-valencia, Jesús A. Rosas-rodríguez, Aldo A. Arvizu-flores, Elisa M. Valenzuela-sotoAbstract:The enzyme betaine aldehyde Dehydrogenase (BADH EC 1.2.1.8) catalyzes the synthesis of glycine betaine (GB), an osmolyte and osmoprotectant. Also, it participates in several metabolic pathways in humans. All BADHs known have cysteine in the active site involved in the aldehyde binding, whereas the porcine kidney enzyme (pkBADH) also has a neighborhood cysteine, both sensitive to oxidation. The antineoplastic and immuno-suppressant pre-drug cyclophosphamide (CTX), and its bioactivation products, have two highly oxidating chlorine atoms. This work aimed to analyze the effect of CTX in the activity of porcine kidney betaine aldehyde Dehydrogenase. PkBADH was incubated with varying CTX concentration (0 to 2.0 mM) at 25 °C and lost 50 % of its activity with 2.0 mM CTX. The presence of the coenzyme NAD^+ (0.5 mM) decreased 95% the activity in 2.0 mM CTX. The substrate betaine aldehyde (0.05 and 0.4 mM, and the products NADH (0.1–0.5 mM) and GB (1 and 10 mM) did not have an effect on the enzyme inactivation by CTX. The reducing agents, dithiothreitol and β-mercaptoethanol, reverted the pkBADH inactivation, but reduced glutathione (GSH) was unable to restore the enzyme activity. Molecular docking showed that CTX could enter at the enzyme active site, where its chlorine atoms may interact with the catalytic and the neighboring cysteines. The results obtained show that CTX inactivates the pkBADH due to oxidation of the catalytic cysteine or because it oxidizes catalytic and neighborhood cysteine, forming a disulfide bridge with a concomitant decrease in the activity of the enzyme.
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Heterogeneity of active sites in recombinant betaine aldehyde Dehydrogenase is modulated by potassium.
Journal of molecular recognition : JMR, 2020Co-Authors: César Muñoz-bacasehua, Jesús A. Rosas-rodríguez, Ciria G. Figueroa-soto, Aldo A. Arvizu-flores, José G. Soñanez-organis, Aurora Stephens‐camacho, Elisa M. Valenzuela-sotoAbstract:Betaine aldehyde Dehydrogenase (BADH EC 1.2.1.8) catalyzes the irreversible oxidation of betaine aldehyde to glycine betaine using NAD+ as a coenzyme. Porcine kidney BADH (pkBADH) follows a bi-bi ordered mechanism in which NAD+ binds to the enzyme before the aldehyde. Previous studies showed that NAD+ induces complex and unusual conformational changes on pkBADH and that potassium is required to maintain its quaternary structure. The aim of this work was to analyze the structural changes in pkBADH caused by NAD+ binding and the role played by potassium in those changes. The pkBADH cDNA was cloned and overexpressed in Escherichia coli, and the protein was purified by affinity chromatography using a chitin matrix. The pkBADH/NAD+ interaction was analyzed by circular dichroism (CD) and by isothermal titration calorimetry (ITC) by titrating the enzyme with NAD+ . The cDNA has an open reading frame of 1485 bp and encodes a protein of 494 amino acids, with a predicted molecular mass of 53.9 kDa. CD data showed that the binding of NAD+ to the enzyme caused changes in its secondary structure, whereas the presence of K+ helps maintain its α-helix content. K+ increased the thermal stability of the pkBADH-NAD+ complex by 5.3°C. ITC data showed that NAD+ binding occurs with different association constants for each active site between 37.5 and 8.6 μM. All the results support previous data in which the enzyme incubation with NAD+ provoked changes in reactivity, which is an indication of slow conformational rearrangements of the active site.
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Inactivation of porcine kidney betaine aldehyde Dehydrogenase by hydrogen peroxide.
Chemico-biological interactions, 2011Co-Authors: Jesús A. Rosas-rodríguez, Elisa M. Valenzuela-sotoAbstract:Concentrated urine formation in the kidney is accompanied by conditions that favor the accumulation of reactive oxygen species (ROS). Under hyperosmotic conditions, medulla cells accumulate glycine betaine, which is an osmolyte synthesized by betaine aldehyde Dehydrogenase (BADH, EC 1.2.1.8). All BADHs identified to date have a highly reactive cysteine residue at the active site, and this cysteine is susceptible to oxidation by hydrogen peroxide. Porcine kidney BADH incubated with H(2)O(2) (0-500 μM) lost 25% of its activity. However, pkBADH inactivation by hydrogen peroxide was limited, even after 120 min of incubation. The presence of coenzyme NAD(+) (10-50 μM) increased the extent of inactivation (60%) at 120 min of reaction, but the ligands betaine aldehyde (50 and 500 μM) and glycine betaine (100 mM) did not change the rate or extent of inactivation as compared to the reaction without ligand. 2-Mercaptoethanol and dithiothreitol, but not reduced glutathione, were able to restore enzyme activity. Mass spectrometry analysis of hydrogen peroxide inactivated BADH revealed oxidation of M278, M243, M241 and H335 in the absence and oxidation of M94, M327 and M278 in the presence of NAD(+). Molecular modeling of BADH revealed that the oxidized methionine and histidine residues are near the NAD(+) binding site. In the presence of the coenzyme, these oxidized residues are proximal to the betaine aldehyde binding site. None of the oxidized amino acid residues participates directly in catalysis. We suggest that pkBADH inactivation by hydrogen peroxide occurs via disulfide bond formation between vicinal catalytic cysteines (C288 and C289).
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Inhibition of porcine kidney betaine aldehyde Dehydrogenase by hydrogen peroxide
Redox report : communications in free radical research, 2010Co-Authors: Jesús A. Rosas-rodríguez, Ciria G. Figueroa-soto, Elisa M. Valenzuela-sotoAbstract:Abstract Renal hyperosmotic conditions may produce reactive oxygen species, which could have a deleterious effect on the enzymes involved in osmoregulation. Hydrogen peroxide was used to provoke oxidative stress in the environment of betaine aldehyde Dehydrogenase in vitro. Enzyme activity was reduced as hydrogen peroxide concentration was increased. Over 50% of the enzyme activity was lost at 100 μM hydrogen peroxide at two temperatures tested. At pH 8.0, under physiological ionic strength conditions, peroxide inhibited the enzyme. Initial velocity assays of betaine aldehyde Dehydrogenase in the presence of hydrogen peroxide (0–200 μM) showed noncompetitive inhibition with respect to NAD+ or to betaine aldehyde at saturating concentrations of the other substrate at pH 7.0 or 8.0. Inhibition data showed that apparent Vmax decreased 40% and 26% under betaine aldehyde and NAD+ saturating concentrations at pH 8.0, while at pH 7.0 Vmax decreased 40% and 29% at betaine aldehyde and NAD+ saturating concentratio...
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Manganese inactivation of renal betaine aldehyde Dehydrogenase from swine
2006Co-Authors: Ciria G. Figueroa-soto, Judith C. Ruíz-lópez, Elisa M. Valenzuela-sotoAbstract:Manganese is an essential micronutrient for mammals, however high manganese concentrations cause adverse health effects. Swine renal betaine aldehyde Dehydrogenase catalyzes the synthesis of glycine betaine, which plays an important role in renal cells osmoregulation. In vitro inactivation of BADH was observed by incubating the purified enzyme in the presence of 1 mM MnCl2 under physiological and low ionic strength conditions. Enzyme inactivation followed first order kinetics in a monophasic process with an inactivation constant of 0.126 ± 0.011 min and 0.137 ± 0.017 at physiological and low ionic strength, respectively. Enzyme inactivation was not prevented by physiological ionic strength, nor by the substrates NAD and betaine aldehyde at saturated concentrations. The enzyme was reactivated with DTT and GSH. Native-PAGE of the inactivated enzyme showed no change in the tetrameric conformation. Intrinsic protein fluorescence studies demonstrated an increased exposure of the tryptophan residues to the aqueous solvent when the enzyme was incubated with Mn. These results suggest that BADH inactivation by Mn may result from the oxidation of cysteines, which induces changes in the tertiary structure of the enzyme.
A.d. Hanson - One of the best experts on this subject based on the ideXlab platform.
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Transgenically Expressed Betaine Aldehyde Dehydrogenase Efficiently Catalyzes Oxidation of Dimethylsulfoniopropionaldehyde and [omega]-Aminoaldehydes
Plant physiology, 1997Co-Authors: Claudine Trossat, Bala Rathinasabapathi, A.d. HansonAbstract:Tobacco (Nicotianum tabacum L.) plants engineered to express a sugar beet (Beta vulgaris L.) betaine aldehyde Dehydrogenase (BADH) cDNA acquired not only BADH activity, but also three other aldehyde Dehydrogenase activities (those measured with 3-dimethylsulfoniopropionaldehyde, 3-aminopropionaldehyde, and 4-aminobutyraldehyde, all of which are natural products). This shows that BADH is not, as believed up to now, a substrate-specific enzyme and that its role may not be limited to glycine betaine synthesis.
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Salt-inducible betaine aldehyde Dehydrogenase from sugar beet: cDNA cloning and expression.
Plant molecular biology, 1992Co-Authors: Kent F. Mccue, A.d. HansonAbstract:Members of the Chenopodiaceae, such as sugar beet and spinach, accumulate glycine betaine in response to salinity or drought stress. The last enzyme in the glycine betaine biosynthetic pathway is betaine aldehyde Dehydrogenase (BADH). In sugar beet the activity of BADH was found to increase two- to four-fold in both leaves and roots as the NaCl level in the irrigation solution was raised from 0 to 500 mM. This increase in BADH activity was paralleled by an increase in level of translatable BADH mRNA. Several cDNAs encoding BADH were cloned from a λgt10 libary representing poly(A)+ RNA from salinized leaves of sugar beet plants, by hybridization with a spinach BADH cDNA. Three nearly full-length cDNA clones were confirmed to encode BADH by their nucleotide and deduced amino acid sequence identity to spinach BADH; these clones showed minor nucleotide sequence differences consistent with their being of two different BADH alleles. The clones averaged 1.7 kb and contained an open reading frame predicting a polypeptide of 500 amino acids with 83% identity to spinach BADH. RNA gel blot analysis of total RNA showed that salinization to 500 mM NaCl increased BADH mRNA levels four-fold in leaves and three-fold in the taproot. DNA gel blot analyses indicated the presence of at least two copies of BADH in the haploid sugar beet genome.
Manabu Ishitani - One of the best experts on this subject based on the ideXlab platform.
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Compatibility of glycinebetaine in rice plants: evaluation using transgenic rice plants with a gene for peroxisomal betaine aldehyde Dehydrogenase from barley
Plant Cell & Environment, 2000Co-Authors: Sachie Kishitani, T. Takanami, M. Suzuki, M. Oikawa, Shuji Yokoi, Manabu Ishitani, A. M. Alvarez‐nakase, Tetsuko TakabeAbstract:Glycinebetaine is synthesized in plants by the two-step oxidation of choline, with betaine aldehyde as the intermediate. The reactions are catalyzed by choline mono-oxygenase and betaine aldehyde Dehydrogenase. Rice plants, which do not accumulate glycinebetaine, possess a gene encoding betaine aldehyde Dehydrogenase, whose activity is detectable at low levels. To evaluate the compatibility in rice of glycinebetaine on growth and tolerance to salt, cold and heat, we produced transgenic rice plants by introduction of a cDNA for betaine aldehyde Dehydrogenase of barley, which is localized in peroxisomes unlike the chloroplast-specific localization of betaine aldehyde Dehydrogenase in spinach and sugar beet. The transgenic rice plants converted high levels of exogenously applied betaine aldehyde (up to 10 mol m 3 ) to glycinebetaine more efficiently than did wild-type plants. The elevated level of glycinebetaine in transgenic plants conferred significant tolerance to salt, cold and heat stress. However, very high levels of glycinebetaine, resulting from conversion of applied betaine aldehyde to glycinebetaine or from exogenous application, inhibited increases in length of rice plants but not increases in dry weight. Our results suggested that the benefits of accumulation of glycinebetaine by rice plants might be considerable under high light conditions.
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Expression of the betaine aldehyde Dehydrogenase gene in barley in response to osmotic stress and abscisic acid.
Plant molecular biology, 1995Co-Authors: Manabu Ishitani, Toshihide Nakamura, Seung Youn Han, Tetsuko TakabeAbstract:When subjected to salt stress or drought, some vascular plants such as barley respond with an increased accumulation of the osmoprotectant glycine betaine (betaine), being the last step of betaine synthesis catalyzed by betaine aldehyde Dehydrogenase (BADH). We report here cloning and characterization of BADH cDNA from barley, a monocot, and the expression pattern of a BADH transcript. An open reading frame of 1515 bp encoded a protein which showed high homology to BADH enzymes present in other plants (spinach and sugar-beet) and in Escherichia coli. Transgenic tobacco plants harboring the clone expressed high levels of both BADH protein and its enzymatic activity. Northern blot analyses indicated that BADH mRNA levels increased almost 8-fold and 2-fold, respectively, in leaves and roots of barley plants grown in high-salt conditions, and that these levels decreased upon release of the stress, whereas they did not decrease under continuous salt stress. BADH transcripts also accumulate in response to water stress or drought, indicating a common response of the plant to osmotic changes that affect its water status. The addition of abscisic acid (ABA) to plants during growth also increased the levels of BADH transcripts dramatically, although the response was delayed when compared to that found for salt-stressed plants. Removal of plant roots before transferring the plants to high-salt conditions reduced only slightly the accumulation of BADH transcripts in the leaves.
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Betaine Aldehyde Dehydrogenase in the Gramineae: Levels in Leaves Both Betaine-Accumulating and Nonaccumulating Cereal Plants
Plant and Cell Physiology, 1993Co-Authors: Manabu Ishitani, Keita Arakawa, Sachie Kishitani, Katsuhiko Mizuno, Tetsuko TakabeAbstract:The accumulation of betaine and the distribution of betaine aldehyde Dehydrogenase, which catalyzes the last step in the synthesis of betaine, were analysed in leaves of control and salt-stressed cereal plants of the Gramineae
Ciria G. Figueroa-soto - One of the best experts on this subject based on the ideXlab platform.
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Heterogeneity of active sites in recombinant betaine aldehyde Dehydrogenase is modulated by potassium.
Journal of molecular recognition : JMR, 2020Co-Authors: César Muñoz-bacasehua, Jesús A. Rosas-rodríguez, Ciria G. Figueroa-soto, Aldo A. Arvizu-flores, José G. Soñanez-organis, Aurora Stephens‐camacho, Elisa M. Valenzuela-sotoAbstract:Betaine aldehyde Dehydrogenase (BADH EC 1.2.1.8) catalyzes the irreversible oxidation of betaine aldehyde to glycine betaine using NAD+ as a coenzyme. Porcine kidney BADH (pkBADH) follows a bi-bi ordered mechanism in which NAD+ binds to the enzyme before the aldehyde. Previous studies showed that NAD+ induces complex and unusual conformational changes on pkBADH and that potassium is required to maintain its quaternary structure. The aim of this work was to analyze the structural changes in pkBADH caused by NAD+ binding and the role played by potassium in those changes. The pkBADH cDNA was cloned and overexpressed in Escherichia coli, and the protein was purified by affinity chromatography using a chitin matrix. The pkBADH/NAD+ interaction was analyzed by circular dichroism (CD) and by isothermal titration calorimetry (ITC) by titrating the enzyme with NAD+ . The cDNA has an open reading frame of 1485 bp and encodes a protein of 494 amino acids, with a predicted molecular mass of 53.9 kDa. CD data showed that the binding of NAD+ to the enzyme caused changes in its secondary structure, whereas the presence of K+ helps maintain its α-helix content. K+ increased the thermal stability of the pkBADH-NAD+ complex by 5.3°C. ITC data showed that NAD+ binding occurs with different association constants for each active site between 37.5 and 8.6 μM. All the results support previous data in which the enzyme incubation with NAD+ provoked changes in reactivity, which is an indication of slow conformational rearrangements of the active site.
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Inhibition of porcine kidney betaine aldehyde Dehydrogenase by hydrogen peroxide
Redox report : communications in free radical research, 2010Co-Authors: Jesús A. Rosas-rodríguez, Ciria G. Figueroa-soto, Elisa M. Valenzuela-sotoAbstract:Abstract Renal hyperosmotic conditions may produce reactive oxygen species, which could have a deleterious effect on the enzymes involved in osmoregulation. Hydrogen peroxide was used to provoke oxidative stress in the environment of betaine aldehyde Dehydrogenase in vitro. Enzyme activity was reduced as hydrogen peroxide concentration was increased. Over 50% of the enzyme activity was lost at 100 μM hydrogen peroxide at two temperatures tested. At pH 8.0, under physiological ionic strength conditions, peroxide inhibited the enzyme. Initial velocity assays of betaine aldehyde Dehydrogenase in the presence of hydrogen peroxide (0–200 μM) showed noncompetitive inhibition with respect to NAD+ or to betaine aldehyde at saturating concentrations of the other substrate at pH 7.0 or 8.0. Inhibition data showed that apparent Vmax decreased 40% and 26% under betaine aldehyde and NAD+ saturating concentrations at pH 8.0, while at pH 7.0 Vmax decreased 40% and 29% at betaine aldehyde and NAD+ saturating concentratio...
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Manganese inactivation of renal betaine aldehyde Dehydrogenase from swine
2006Co-Authors: Ciria G. Figueroa-soto, Judith C. Ruíz-lópez, Elisa M. Valenzuela-sotoAbstract:Manganese is an essential micronutrient for mammals, however high manganese concentrations cause adverse health effects. Swine renal betaine aldehyde Dehydrogenase catalyzes the synthesis of glycine betaine, which plays an important role in renal cells osmoregulation. In vitro inactivation of BADH was observed by incubating the purified enzyme in the presence of 1 mM MnCl2 under physiological and low ionic strength conditions. Enzyme inactivation followed first order kinetics in a monophasic process with an inactivation constant of 0.126 ± 0.011 min and 0.137 ± 0.017 at physiological and low ionic strength, respectively. Enzyme inactivation was not prevented by physiological ionic strength, nor by the substrates NAD and betaine aldehyde at saturated concentrations. The enzyme was reactivated with DTT and GSH. Native-PAGE of the inactivated enzyme showed no change in the tetrameric conformation. Intrinsic protein fluorescence studies demonstrated an increased exposure of the tryptophan residues to the aqueous solvent when the enzyme was incubated with Mn. These results suggest that BADH inactivation by Mn may result from the oxidation of cysteines, which induces changes in the tertiary structure of the enzyme.
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Original article: Manganese inactivation of renal betaine aldehyde Dehydrogenase from swine
2006Co-Authors: Judith C. Ruíz-lópez, Ciria G. Figueroa-soto, Elisa M. Valenzuela-sotoAbstract:Manganese is an essential micronutrient for mammals, however high manganese concentrations cause adverse health effects. Swine renal betaine aldehyde Dehydrogenase catalyzes the synthesis of glycine betaine, which plays an important role in renal cells osmoregulation. In vitro inactivation of BADH was observed by incubating the purified enzyme in the presence of 1 mM MnCl2 under physiological and low ionic strength conditions. Enzyme inactivation followed first order kinetics in a monophasic process with an inactivation constant of 0.126 ± 0.011 min -1 and 0.137 ± 0.017 at physiological and low ionic strength, respectively. Enzyme inactivation was not prevented by physiological ionic strength, nor by the substrates NAD + and betaine aldehyde at saturated concentrations. The enzyme was reactivated with DTT and GSH. Native-PAGE of the inactivated enzyme showed no change in the tetrameric conformation. Intrinsic protein fluorescence studies demonstrated an increased exposure of the tryptophan residues to the aqueous solvent when the enzyme was incubated with Mn 2+ . These results suggest that BADH inactivation by Mn 2+ may result from the oxidation of cysteines, which induces changes in the tertiary structure of the enzyme.
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Purification of a heterodimeric betaine aldehyde Dehydrogenase from wild amaranth plants subjected to water deficit.
Biochemical and biophysical research communications, 2001Co-Authors: Ciria G. Figueroa-soto, Elisa M. Valenzuela-sotoAbstract:Abstract Betaine aldehyde Dehydrogenase was purified to homogeneity from wild-type amaranth plants subjected to water deficit. The enzyme has a native molecular mass of 125 kDa; it is formed by two subunits, one of the subunits with a molecular mass of 63 kDa and the second one of 70 kDa as determined by SDS–PAGE and double dimension electrophoresis. IEF studies showed two bands with pI values of 4.93 and 4.85, respectively. Possible glycosilation of the 63- and 70-kDa subunits were tested with negative results. Both subunits cross-reacted strongly with polyclonal antibody raised against porcine kidney BADH. Also antiserum rose against HSP70 cross-reacted strongly with the wild amaranth BADH 70-kDa subunit. The enzyme was stable to extreme pH's and temperatures, and high KCl concentrations. Product inhibition of BADH was not observed.