Betaine Aldehyde Dehydrogenase Activity Research Articles

Comparative Effects of Salt Stress and Extreme pH Stress Combined on Glycinebetaine Accumulation, Photosynthetic Abilities and Growth Characters of Two Rice Genotypes

Glycinebetaine (Glybet) accumulation, photosynthetic efficiency and growth performance in indica rice cultivated under salt stress and extreme pH stress were investigated. Betaine aldehyde dehydrogenase (BADH) activity and Glybet accumulation in the seedlings of salt-tolerant and salt-sensitive rice varieties grown under saline and acidic conditions peaked after treatment for 72 h and 96 h, respectively, and were higher than those grown under neutral pH and alkaline salt stress. A positive correlation was found between BADH activity and Glybet content in both salt-tolerant ( r 2 = 0.71) and salt-sensitive ( r 2 = 0.86) genotypes. The chlorophyll a, chlorophyll b, total chlorophyll and total carotenoids contents in the stressed seedlings significantly decreased under both acidic and alkaline stresses, especially in the salt-sensitive genotype. Similarly, the maximum quantum yield of PSII ( F v/ F m), photon yield of PSII (Φ PSII), non-photochemical quenching (NPQ) and net photosynthetic rate ( P n) in the stressed seedlings were inhibited, leading to overall growth reduction. The positive correlations between chlorophyll a content and F v/ F m, total chlorophyll content and Φ PSII, Φ PSII and P n as well as P n and leaf area in both salt-tolerant and salt-sensitive genotypes were found. Saline acidic and saline alkaline soils may play a key role affecting vegetative growth prior to the reproductive stage in rice plants.

Drought and Salinity Tolerance in Transgenic Potato Expressing the Betaine Aldehyde Dehydrogenase Gene

Glycine betaine (GB) is a common compatible solute in many different organisms including higher plants. Many plant species can accumulate GB in response to drought and salinity. GB is synthesized by conversion of choline to GB through a two-step oxidation via the intermadiate betaine aldehyde. In higher plants, the relevant enzymes are choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BADH). The fact that many important crops, such as rice, potato and tomato are betaine-deficient has inevitably led to the proposal that it might be possible to increase drought and salinity tolerances by genetic engineering of GB synthesis. In the present study, the transgenic plants of potato cultivar Gannongshu 2 were obtained by Agro- bacterium-mediated transformation of the expression vector pBIBB contained BADH gene under the control of the constitutive promoter CaMV 35S. PCR, Southern, and Northern blot analyses showed that the BADH gene was integrated into potato genome, transcribed and expressed in the transgenic plants. The analysis of BADH activity of transgenic plant leaves revealed that the BADH activity ranged from 2.1 to 10.5 U, while it was not detectable in the control plants. There was a negative relationship (y = –3.774x+57.083, r = 0.989**) between BADH activity and relative electric conductivity of the transgenic potato leaves. The trans- genic potato plants grew normally under NaCl and polyethylene glycol (PEG) stresses with increase of 0.41–1.00 cm for plant height and 10–35% for fresh weight per plant compared with the control plants. This result demonstrated that the transgenic potato plants can improve tolerances to drought and salinity as a result of transformation and expression of BADH gene.

Physiological and Growth Responses of Tomato Progenies Harboring the Betaine Alhyde Dehydrogenase Gene to Salt Stress

AbstractThe responses of five transgenic tomato (Lycopersicon esculentum Mill) lines containing the betaine aldehyde dehydrogenase (BADH) gene to salt stress were evaluated. Proline, betaine (N, N, N‐trimethylglycine, hereafter betaine), chlorophyll and ion contents, BADH activity, electrolyte leakage (EL), and some growth parameters of the plants under 1.0% and 1.5% NaCl treatments were examined. The transgenic tomatoes had enhanced BADH activity and betaine content, compared to the wild type under stress conditions. Salt stress reduced chlorophyll contents to a higher extent in the wild type than in the transgenic plants. The wild type exhibited significantly higher proline content than the transgenic plants at 0.9% and 1.3% NaCl. Cell membrane of the wild type was seaerely damaged as determined by higher EL under salinity stress. K+and Ca2+ contents of all tested lines decreased under salt stress, but the transgenic plants showed a significantly higher accumulation of K+ and Ca2+ than the wild type. In contrast, the wild type had significantly higher Cl and Na+ contents than the transgenic plants under salt stress. Although yield reduction among various lines varied, the wild type had the highest yield reduction. Fruit quality of the transgenic plants was better in comparison with the wild type as shown by a low ratio of blossom end rot fruits. The results show that the transgenic plants have improved salt tolerance over the wild type.(Handling editor: Jin‐Zhong Cui)

Glycinebetaine Accumulation, Physiological Characterizations and Growth Efficiency in Salt‐tolerant and Salt‐sensitive Lines of Indica Rice (Oryza sativaL. ssp.indica) in Response to Salt Stress

AbstractThe aim of this research was to investigate betaine aldehyde dehydrogenase (BADH) and glycinebetaine (Glybet) biosynthesis in photoautotrophic rice seedlings. The role of Glybet on physiological and growth responses to salt stress in both salt‐tolerant and salt‐sensitive lines is to be investigated. The BADH activity in salt‐tolerant seedlings cultured under extreme salt stress (342 mmNaCl) progressively increased during the first few hours until it peaked after 72 h. This was about 2.5 times greater than in salt‐sensitive plants. Similarly, the amount of Glybet detected in salt‐tolerant lines was 1.3 times more than in salt‐sensitive lines at 96 h salt exposure. The BADH activities were positively related to Glybet accumulation in both salt‐tolerant and salt‐sensitive lines. The accumulation of Glybet in salt‐tolerant lines was directly correlated with pigment stabilization. Relative water content in the salt‐tolerant lines was closely related to water oxidation in photosystem II (PSII), defined by maximum quantum yield of PSII (Fv/Fm). In addition, a high concentration of total chlorophyll is more efficient in capturing light energy, defined by photochemical quenching. The concentrations of chlorophyllaand total carotenoid were positively related to the quantum efficiency of PSII (ΦPSII) and non‐photochemical quenching, respectively, resulting in a high net‐photosynthetic rate (NPR) and the promotion of growth. The high level of Glybet in salt‐tolerant lines plays a role as a salt defensive response mechanism in terms of pigment stabilization and water oxidation in PSII, resulting in high NPR and growth efficiency.

The accumulation and synthesis of betaine in winter skate (Leucoraja ocellata)

The present study investigated aspects of betaine metabolism in an elasmobranch fish, the winter skate (Leucoraja ocellata). Based on the level of choline dehydrogenase (ChoDH) activity, the liver and kidney appear to be the major sites of betaine synthesis and the mitochondrial localization of ChoDH and betaine aldehyde dehydrogenase (BADH) indicates that the metabolic organization of betaine synthesis in winter skate is similar to other vertebrates. Food deprivation did not affect white muscle betaine content, and prolonged starvation (70 days) appeared to decrease the total hepatic betaine synthetic capacity. There was no decrease in ChoDH or BADH activity at the mitochondrial level with starvation, suggesting any decrease is due to catabolism of hepatic reserves rather than downregulation of betaine synthesis. Skates fed a high betaine diet (frozen squid approximately 55 micromol g(-1)) had elevated white muscle betaine content compared to those fed a low betaine diet (frozen herring <2 micromol g(-1)); however, high dietary betaine intake did not affect the activity of betaine synthesizing enzymes in liver. Acclimation to elevated salinity (120 and 130% seawater) did not result in an increase in white muscle betaine content. Taken as a whole, the present data suggest that diet is a major determinant of muscle betaine in the winter skate and that betaine is of marginal importance as an intracellular osmolyte in this species.

Betaine aldehyde dehydrogenase from Pseudomonas aeruginosa: cloning, over-expression in Escherichia coli, and regulation by choline and salt

In the human pathogen Pseudomonas aeruginosa, betaine aldehyde dehydrogenase (BADH) may play a dual role assimilating carbon and nitrogen from choline or choline precursors--abundant at infection sites--and producing glycine betaine, which protects the bacteria against the high-osmolarity stress prevalent in the infected tissues. We cloned the P. aeruginosa BADH gene and expressed the BADH protein in Escherichia coli. The recombinant protein appears identical to its native counterpart, as judged by Western blot, N-terminal amino acid sequence, tryptophan-fluorescence emission spectra, circular-dichroism spectroscopy, size-exclusion chromatography, and kinetic properties. Computational analysis indicated that the promoter sequence of the putative operon that includes the BADH gene has a consensus-binding site for the choline-sensing transcription repressor BetI, and putative boxes for ArcA and Lrp transcription factors but no known elements of response to osmotic stress. This is consistent with the strong induction of BADH expression by choline and with the lack of effect of NaCl. As there were significant amounts of BADH protein and activity in P. aeruginosa cells grown on glucose plus choline, as well as the BADH activity exhibiting tolerance to salt, it is likely that glycine betaine is synthesized in vivo and could play an important osmoprotectant role under conditions of infection.

Inactivation of betaine aldehyde dehydrogenase from Pseudomonas aeruginosa and Amaranthus hypochondriacus L. leaves by disulfiram

Betaine aldehyde dehydrogenase (BADH) activity might be crucial for the growth of the human pathogen Pseudomonas aeruginosa under conditions of infection and therefore appears to be a suitable target for antimicrobial agents. As a first step in the search for BADH inhibitors, we have tested the effects of the known aldehyde dehydrogenase inhibitor disulfiram (DSF) on the activity of P. aeruginosa and Amaranthus hypochondriacus (amaranth) leaf BADHs. DSF totally inactivated both enzymes in a time- and dose-dependent manner. In the case of the Pseudomonas enzyme, inactivation kinetics were monophasic with a second-order inactivation rate constant at pH 6.9 of 4.9±0.4 M −1 s −1, whereas the plant enzyme was inactivated in a biphasic process with second-order inactivation rate constants at pH 7.5 of 6.8±0.6 and 0.33±0.04 M −1 s −1. At pH 8.8, the second-order rate constants for inactivation of the bacterial enzyme was 1×10 3 M −1 s −1, which compare well with that reported for human liver mitochondrial aldehyde dehydrogenase (ALDH2), the target of DSF inhibition in the aversion therapy of alcoholism. Both BADHs were inactivated faster in the presence of NAD(P) + than in its absence, whereas NAD(P)H and betaine aldehyde protected the bacterial, but increased the inactivation rate of the plant enzyme. The inactivated enzymes were reactivated by dithiothreitol, but not by a high concentration of the physiological reductant glutathione. The high in vitro sensitivity of the Pseudomonas BADH to DSF, particularly in the presence of NAD(P) +, together with the lack of reversibility of DSF modification by glutathione, makes this inhibitor a potential antimicrobial agent and suggests that it might be worth testing its effects and those of its metabolites in vivo, under culture conditions in which the activity of BADH is required for growth of the bacteria.

Betaine aldehyde dehydrogenase kinetics partially account for oyster population differences in glycine betaine synthesis

Betaine aldehyde dehydrogenase (BADH), the terminal enzyme of the glycine betaine synthetic pathway was purified 245-fold from the mitochondria of Atlantic and Chesapeake Bay oyster populations acclimated to 350 mosm, using ammonium sulfate precipitation, anion exchange, and affinity chromatography. BADH from both populations functions at its maximum rate at 50-55 degrees C over a broad pH range (7.5-9). BADH activity is also modulated by increased [Na(+)] and [K(+)]. Although BADH from both populations has a similar V(max), BADH from Bay oysters has a substantially lower affinity for its substrate, betaine aldehyde, (K(m) = 0.36 mM), than BADH from Atlantic oysters (K(m) = 0.1 mM). Despite kinetic differences, BADH from both Atlantic and Chesapeake Bay oysters have the same molecular weight based on electrophoretic analysis. These differences in BADH enzyme kinetics between the two oyster populations probably partially explain the lower glycine betaine synthesis rates and concentrations in Chesapeake Bay oysters. J. Exp. Zool. 286:238-249, 2000.

Occurrence of choline and glycine betaine uptake and metabolism in the family rhizobiaceae and their roles in osmoprotection

The role of glycine betaine and choline in osmoprotection of various Rhizobium, Sinorhizobium, Mesorhizobium, Agrobacterium, and Bradyrhizobium reference strains which display a large variation in salt tolerance was investigated. When externally provided, both compounds enhanced the growth of Rhizobium tropici, Sinorhizobium meliloti, Sinorhizobium fredii, Rhizobium galegae, Agrobacterium tumefaciens, Mesorhizobium loti, and Mesorhizobium huakuii, demonstrating their utilization as osmoprotectants. However, both compounds were inefficient for the most salt-sensitive strains, such as Rhizobium leguminosarum (all biovars), Agrobacterium rhizogenes, Rhizobium etli, and Bradyrhizobium japonicum. Except for B. japonicum, all strains exhibit transport activity for glycine betaine and choline. When the medium osmolarity was raised, choline uptake activity was inhibited, whereas glycine betaine uptake was either increased in R. leguminosarum and S. meliloti or, more surprisingly, reduced in R. tropici, S. fredii, and M. loti. The transport of glycine betaine was increased by growing the cells in the presence of the substrate. With the exception of B. japonicum, all strains were able to use glycine betaine and choline as sole carbon and nitrogen sources. This catabolic function, reported for only a few soil bacteria, could increase competitiveness of rhizobial species in the rhizosphere. Choline dehydrogenase and betaine-aldehyde dehydrogenase activities were present in the cells of all strains with the exception of M. huakuii and B. japonicum. The main physiological role of glycine betaine in the family Rhizobiaceae seems to be as an energy source, while its contribution to osmoprotection is restricted to certain strains.

Rapid purification and properties of betaine aldehyde dehydrogenase from Pseudomonas aeruginosa.

Betaine aldehyde dehydrogenase (BADH) (EC 1.2.1.8) catalyzes the last, irreversible step in the synthesis of the osmoprotectant glycine betaine from choline. In Pseudomonas aeruginosa this reaction is also an obligatory step in the assimilation of carbon and nitrogen when bacteria are growing in choline or choline precursors. We present here a method for the rapid purification to homogeneity of this enzyme by the use of ion-exchange and affinity chromatographies on 2',5'-ADP-Sepharose, which results in a high yield of pure enzyme with a specific activity at 30 degreesC and pH 7.4 of 74.5 U/mg of protein. Analytical ultracentrifugation, gel filtration, chemical cross-linking, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggest that BADH from P. aeruginosa is a homodimer with 61-kDa subunits. The amino acid composition and the N-terminal sequence of 21 amino acid residues showed significant similarity with those of the enzymes from Xanthomonas translucens and Escherichia coli. Neither BADH activity nor BADH protein was found in cell extracts from bacteria grown in the absence of choline. In contrast to other BADHs studied to date, the Pseudomonas enzyme cannot use positively charged aldehydes other than betaine aldehyde as substrates. The oxidation reaction has an activation energy of 39.8 kJ mol-1. The pH dependence of the velocity indicated an optimum at pH 8.0 to 8.5 and the existence of two ionizable groups with macroscopic pK values of 7.0 +/- 0.1 and 9. 7 +/- 0.1 involved in catalysis and/or binding of substrates. The enzyme is inactivated at 40 degreesC, but activity is regained when the heated enzyme is cooled to 30 degreesC or lower. At the optimum pH of 8.0, the enzyme is inactivated by dilution, but it is stable at pH 6.5 even at very low concentrations. Also, P. aeruginosa BADH activity is rapidly lost on removal of K+. In all cases studied, inactivation involves a biphasic process, which was dependent on the enzyme concentration only in the case of inactivation by dilution. NADP+ considerably protected the enzyme against these inactivating conditions.

Accumulation of glycinebetaine and its synthesis from radioactive precursors under salt-stress in the cyanobacterium Aphanothece halophytica

Growth in salt-stressed (2.0 M NaCl) Aphanothece halophytica was initially delayed during the first two days of cultivation and eventually attained the same growth rate as the control (0.5 M NaCl) cells. Glycinebetaine accumulation increased slightly in control cells but a dramatic increase of glycinebetaine occurred in salt-stressed cells during a growth period of six days. There was no apparent increase in the synthesis of [14C] glycinebetaine in the control cells, in contrast to the marked increase in its synthesis in the salt-stressed cells. Increasing NaCl concentration in the growth medium induced both the accumulation and the synthesis of glycinebetaine. Time course experiments provided evidence that [14C] choline was first oxidized to [14C] betaine aldehyde which was further oxidized to [14C] glycinebetaine in A. halophytica. The supporting data for such a pathway were obtained from the presence of choline and betaine aldehyde dehydrogenase activities found in the membrane and cytoplasmic fractions, respectively. The activities of these two enzymes were also enhanced upon increasing NaCl concentration in the growth medium from 0.5 M to 2.0 M. Under this condition an increaseof approximately 1.5-fold was observed for choline dehydrogenase activity as compared to 2.5-fold for betaine aldehyde dehydrogenase activity, suggesting a preferable induction of the latter enzyme by salt stress. A. halophytica was able to utilize [14C] ethanolamine and [14C] glycine for the synthesis of [14C] glycinebetaine.

Porcine kidney betaine aldehyde dehydrogenase: purification and properties

Significant betaine aldehyde dehydrogenase activity was found in porcine kidney. The enzyme was purified 320-fold with an overall recovery of 11%. It had a specific activity of 115.8 nkats/mg protein and proved to be homogeneous by SDS-PAGE with a subunit molecular mass of 52 kDa. IEF studies showed three bands with pI values of 5.74, 5.68 and 5.58, respectively. The enzyme was stable in a pH range between 5.0 and 10.0 and the optimum pH was 9.5. The reaction is highly specific for NAD + and betaine aldehyde, although acetaldehyde, butyraldehyde and glyceraldehyde can be used. Estimated values of K m at pH 8.0 and 25°C were 127 μM for betaine aldehyde and 40 μM for NAD +. The reaction could not be reversed even at high glycine betaine concentrations. The enzyme was not activated by salts at high concentrations but it was salt tolerant—retaining 50% of maximal activity at 1.0 M K + and Na +. It is inferred that salt tolerance is an essential property for an enzyme participating in the cellular synthesis of an osmoprotectant. Proline, glycerol, sucrose and mannitol had a little effect on the enzyme activity while glycine betaine had an inhibitory effect.

Expression of betaine aldehyde dehydrogenase gene and salinity tolerance in rice transgenic plants

Betaine as one of osmolytes plays an important role in osmoregulation of most high plants. Betaine aldehyde dehydrogenase (BADH) is the second enzyme involved in betaine biosynthesis. The BADH gene from a halophite,Atriplex hortensis, was transformed into rice cultivars by bombarment method. Totally 192 transgenic rice plants were obtained and most of them had higher salt tolerance than controls. Among transgenic plants transplanted in the saline pool containing 0.5% NaCl in a greenhouse, 22 survived. 13 of which set seeds, and the frequency of seed setting was very low, only 10%. But the controls could not grow under the same condition. The results of BADH activity assay and Northern blot showed that the BADH gene was integrated into chromosomes of transgenic plants and expressed.

Transgenically Expressed Betaine Aldehyde Dehydrogenase Efficiently Catalyzes Oxidation of Dimethylsulfoniopropionaldehyde and [omega]-Aminoaldehydes.

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.

Possible Involvement of Greening in Cell Growth of Suaeda japonica under Salt Stress

Suaeda japonica, a member of the family Chenopodiaceae, is a halophyte that grows on the shores of the Ariake Sea in Japan. Using yellowish and selected green callus tissues of S. japonica, we examined the correlation between cell growth and glycinebetaine content, as well as the activity of betaine aldehyde dehydrogenase (BADH), under salt stress. Although the growth of yellowish callus tissues was markedly inhibited by 0.1 M NaCl, that of green callus tissues was not. However, in 0.3 M NaCl, the fresh weight of green callus tissues was decreased by 15%. Both the endogenous level of glycinebetaine and the activity of BADH in yellowish callus tissues were very low, but green callus tissues contained very high levels of glycinebetaine and high BADH activity. The activity in green callus increased significantly with increases in the concentration of NaCl. The increase in levels of Chl was also observed, but the increase occurred earlier than that in the level of glycinebetaine. Functional development of chloroplasts (an increase in the level of Chl) seems to be involved in the increase in the activity of BADH, and the resultant increase in levels of glycinebetaine seems to facilitate callus growth under salt stress.

Molecular characterization of mutants affected in the osmoprotectant-dependent induction of phospholipase C in Pseudomonas aeruginosa PAO1.

Production of the two phospholipases C (PLCs) in Pseudomonas aeruginosa PAO1 is induced under conditions of phosphate limitation, or by the osmoprotectants choline or glycine betaine. Tn5 mutagenesis was performed on strain PAO1 to isolate mutants deficient in choline-dependent induction of PLC. Two mutants, Tn5T1 and Tn5G19, were identified which produce decreased levels of PLC in phosphate-replete media supplemented with choline. A total of 136 and 496 bp of flanking DNA from Tn5G19 and Tn5T1 was cloned by an inverse polymerase chain reaction (PCR) and sequenced. The DNA flanking the Tn5T1 insertion contains an open reading frame predicted to encode a peptide that is approx. 60% identical to the N-terminus of a previously identified protein (P35) of unknown function from Escherichia coli. The P35 gene, which is located in the nusA-infB operon in E. coll, was designated orp (osmoprotectant regulator of PLC). Haemolytic titres, total PlcH protein and beta-galactosidase activity expressed from a chromosomally inserted plcH-lacZ operon fusion were reduced in strain Tn5T1 in comparison with the parental strain (PAO1) carrying the same fusion. However, this mutant expressed several-fold higher levels of plcH message than strain PAO1 in the presence of choline, while the phosphate-starvation-dependent transcript of plcH could not be detected in this mutant. The defects in Tn5T1 are complemented by a DNA fragment, isolated from a genomic library of PAO1, that carries the orp gene. The deduced amino acid sequence of the DNA fragment cloned from Tn5G19 exhibits 84% identity with the betB gene product of E. coli that has betaine aldehyde dehydrogenase activity. This enzyme catalyses the conversion of betaine aldehyde to glycine-betaine. Unlike the parental strain, the Tn5G19 mutant could not utilize choline as a sole carbon, nitrogen and energy source, and it was deficient in betaine aldehyde dehydrogenase activity. Also, consistent with a disruption of betB in Tn5G19, choline inhibited growth of this strain in media containing 0.7 M NaCl, while glycine-betaine restores growth to wild-type levels. The defects in Tn5G19 are complemented by a DNA fragment from PAO1 that carries the betB gene. The orp gene is located between 0.6 to 6.6 min while betB is located between 10.5 to 12.5 min on the chromosome of PAO1.

Human Aldehyde Dehydrogenase E3 Isozyme Is a Betaine Aldehyde Dehydrogenase

The E3 isozyme of human aldehyde dehydrogenase (EC 1.2.1.3), with broad substrate specificity, which also catalyzes dehydrogenation of 4-aminobutyraldehyde, was purified and sequenced recently (1,3). It has been shown during this investigation to have betaine aldehyde dehydrogenase activity. Betaine aldehyde and 4-aminobutyraldehyde activities copurified on six chromatographic columns. Molecular properties of the homogeneous product were identical with those of E3 isozyme. Activity with betaine aldehyde was considerably higher than that with 4-aminobutyraldehyde, the best known substrate. Thus, human E3 isozyme and betaine aldehyde dehydrogenase (EC 1.2.1.8) are the same enzyme.

Characterization of partially purified betaine aldehyde dehydrogenase from horseshoe crab (Limulus polyphemus) cardiac mitochondria

AbstractGlycine betaine is an organic osmolyte which is often accumulated in response to hyperosmotic stress in euryhaline species. In mitochondria isolated from horseshoe crab (Limulus polyphemus) heart tissue, the synthetic pathway, which is a two step oxidation of choline (choline→betaine aldehyde→glycine betaine), appears to be associated with the matrix. The majority of the betaine aldehyde dehydrogenase (BADH) activity, the terminal synthetic enzyme, is found in this subcellular fraction. Partially purified Limulus BADH is highly specific for the substrate betaine aldehyde (Km of 133 μM). Two alternative substrates, glyceraldehyde and glycoaldehyde, do not stimulate NAD+ reduction in the presence of BADH. NAD+ is an essential co‐factor in the oxidation of betaine aldehyde to glycine betaine. Limulus BADH can utilize NADP+, although a higher Km indicates a much lower affinity than that for NAD+, 267 μM and 22 μM, respectively. The end‐product, glycine betaine, competitively inhibits BADH activity. Cations have a profound effect on Limulus BADH activity. At physiological concentrations, enzyme activity is stimulated by increases in both [K+] but decreased by elevated [Na+]. Although inorganic ions modulate this enzyme, it does not appear that BADH is the rate limiting step in osmotically stimulated glycine betaine synthesis. © Wiley‐Liss, Inc.

Seed germination of the halophyteSuaeda japonica under salt stress

Suaeda japonica Makino belonging to the family Chenopodiaceae, is a halophyte and grows at the shore of Ariake sea in Japan. This plant presumably possesses high salt resistant nature, thus, we examined the mechanisms of seed germination under salt stress. The seeds maintained 80% germination rates on the medium containing 0.7 M NaCl. Germination rates varied depending on salt type; the germination rates under NaCl or KCI exhibited relatively lower values than ones under sodium gluconate or potassium gluconate. This different responses for salts seemed to be as a result of the presence of Cl− ions. Although very high levels of betaine (compatible solute), were kept in the seedlings grown under no salt stress, the contents gradually increased as concentration of NaCl increased. Betaine is a factor present in plants that works to alleviate the effects of excessive soil salts. It is synthesized in leaves from betaine aldehyde, and this process is catabolized by betaine aldehyde dehydrogenase (BADH). When the seedlings were cultivated on the medium without NaCl, relatively high level of BADH activity was found. The activity increased 5-fold in the seedlings grown under 0.5 M NaCl stress. Increases in betaine content and BADH activity were found during seed germination. InS. japonica, the salt stress promoted BADH activity, subsequently endogenous betaine contents were increased, and increased betaine seemed to secure seed germination under salt stress.

Salt-inducible betaine aldehyde dehydrogenase from sugar beet: cDNA cloning and expression

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 lambda gt10 library 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.