The soybean aldehyde dehydrogenase (ALDH) protein superfamily

Aldehyde dehydrogenases (ALDHs) are members of NAD(P)(+)-dependent protein superfamily that catalyze the oxidation of a wide range of endogenous and exogenous highly reactive aliphatic and aromatic aldehyde molecules to their corresponding non toxic carboxylic acids. Research evidence has shown that ALDHs represent a promising class of genes to improve growth development, seed storage and environmental stress adaptation in higher plants. The recently completed genome sequences of several plant species have resulted in the identification of a large number of ALDH genes, most of which still need to be functionally characterized. In this paper, we identify members of the ALDH gene superfamily in soybean genome, and provide a unified nomenclature for the entire soybean ALDH gene families. The soybean genome contains 18 unique ALDH sequences encoding members of five ALDH families involved in a wide range of metabolic and molecular detoxification pathways. In addition, we describe the biochemical requirements and cellular metabolic pathways of selected members of ALDHs in soybean responses to environmental stress conditions.     

Inhibition of cytosolic class 3 aldehyde dehydrogenase by antisense oligonucleotides in rat hepatoma cells

Aldehyde dehydrogenases (ALDHs) are a superfamily of several isoenzymes widely expressed in bacteria, yeast, plant and animals. Three major classes of ALDHs have been traditionally identified, classes 1, 2 and 3. Both exogenous and endogenous aldehydes, including aldehydes derived from lipid peroxidation, are oxidized by the ALDH superfamily. Several changes in ALDH isoenzyme expression take place in hepatoma cells, in particular cytosolic class 3 ALDH (ALDH3), not expressed in normal hepatocytes, appears and increases with the degree of deviation. It has been demonstrated that cytosolic ALDH3 is important in determining the resistance of tumor cells to antitumor drugs, such as cyclophosphamide. Moreover, hepatoma-associated ALDH3 seems to be important in metabolizing aldehydes derived from lipid peroxidation, and in particular the cytostatic aldehyde 4-hydroxynonenal (4-HNE). We demonstrated previously that restoring endogenous lipid peroxidation in hepatoma cells by enriching them with arachidonic acid causes a decrease of mRNA, protein and enzyme activity of ALDH3 and that this decrease reduces cell growth and/or causes cell death, depending on basal class 3 ALDH activity. To confirm the correlation between inhibition of class 3 ALDH and reduction of cell proliferation, we exposed hepatoma cells to antisense oligonucleotides (ODNs) against ALDH3. In JM2 hepatoma cell line, with high ALDH3 activity, the exposure to antisense ODNs significantly decreases mRNA and enzyme activity (90%). At the same time, cell growth was reduced by about 70%. The results confirm that in hepatoma cells ALDH3 expression is closely related with cell growth, and that its inhibition is important in reducing the proliferation of hepatoma cells overexpressing ALDH3.     

Aldehyde Dehydrogenases and Their Role in Carcinogenesis

Abstract

Aldehydes are highly reactive molecules that may have a variety of effects on biological systems. They can be generated from a virtually limitless number of endogenous and exogenous sources. Although some aldehyde-mediated effects such as vision are beneficial, many effects are deleterious, including cytotoxicity, mutagenicity, and carcinogenicity. A variety of enzymes have evolved to metabolize aldehydes to less reactive forms. Among the most effective pathways for aldehyde metabolism is their oxidation to carboxylic acids by aldehyde dehydrogenases (ALDHs).

ALDHs are a family of NADP-dependent enzymes with common structural and functional features that catalyze the oxidation of a broad spectrum of aliphatic and aromatic aldehydes. Based on primary sequence analysis, three major classes of mammalian ALDHs — 1, 2, and 3 — have been identified. Classes 1 and 3 contain both constitutively expressed and inducible cytosolic forms. Class 2 consists of constitutive mitochondrial enzymes. Each class appears to oxidize a variety of substrates that may be derived either from endogenous sources such as amino acid, biogenic amine, or lipid metabolism or from exogenous sources, including aldehydes derived from xenobiotic metabolism.

Changes in ALDH activity have been observed during experimental liver and urinary bladder carcinogenesis and in a number of human tumors, including some liver, colon, and mammary cancers. Changes in ALDH define at least one population of preneoplastic cells having a high probability of progressing to overt neoplasms. The most common change is the appearance of class 3 ALDH dehydrogenase activity in tumors arising in tissues that normally do not express this form. The changes in enzyme activity occur early in tumorigenesis and are the result of permanent changes in ALDH gene expression.

This review discusses several aspects of ALDH expression during carcinogenesis. A brief introduction examines the variety of sources of aldehydes. This is followed by a discussion of the mammalian ALDHs. Because the ALDHs are a relatively understudied family of enzymes, this section presents what is currently known about the general structural and functional properties of the enzymes and the interrelationships of the various forms.

The remainder of the review discusses various aspects of the ALDHs in relation to tumorigenesis. The expression of ALDH during experimental carcinogenesis and what is known about the molecular mechanisms underlying those changes are discussed. This is followed by an extended discussion of the potential roles for ALDH in tumorigenesis. The role of ALDH in the metabolism of cyclophosphamidelike chemotherapeutic agents is described. This work suggests that modulation of ALDH activity may be an important determinant of the effectiveness of certain chemotherapeutic agents. The evidence that changes in ALDH are part of an adaptive response of preneoplastic and neoplastic cells to altered cell physiology or stress is then considered. Roles in the metabolism of aldehydes generated from lipid peroxidation and as part of the Ah gene-mediated response to xenobiotic exposure are both discussed. The data are consistent with a role for certain ALDHs in lipid aldehyde metabolism. Biochemical and genetic data also imply that changes in ALDH may be linked, in part, to cellular adaptation to oxidative stress.

Finally, a model of inducible ALDH gene regulation is proposed. The model incorporates current information about ALDH gene expression with the regulation of other genes known to be part of the adaptive responses occurring in neoplastic cells. The model suggests that regulation of class 1 and 3 ALDH gene activity may be complex, involving the tissue-specific ability to respond to a variety of physiological cues. The model also suggests several avenues for future research that should provide a clearer understanding of the regulation of this important gene family in response to a variety of factors.     

Inhibition of cytosolic class 3 aldehyde dehydrogenase by antisense oligonucleotides in rat hepatoma cells

Aldehyde dehydrogenases (ALDHs) are a superfamily of several isoenzymes widely expressed in bacteria, yeast, plant and animals. Three major classes of ALDHs have been traditionally identified, classes 1, 2 and 3. Both exogenous and endogenous aldehydes, including aldehydes derived from lipid peroxidation, are oxidized by the ALDH superfamily. Several changes in ALDH isoenzyme expression take place in hepatoma cells, in particular cytosolic class 3 ALDH (ALDH3), not expressed in normal hepatocytes, appears and increases with the degree of deviation. It has been demonstrated that cytosolic ALDH3 is important in determining the resistance of tumor cells to antitumor drugs, such as cyclophosphamide. Moreover, hepatoma-associated ALDH3 seems to be important in metabolizing aldehydes derived from lipid peroxidation, and in particular the cytostatic aldehyde 4-hydroxynonenal (4-HNE). We demonstrated previously that restoring endogenous lipid peroxidation in hepatoma cells by enriching them with arachidonic acid causes a decrease of mRNA, protein and enzyme activity of ALDH3 and that this decrease reduces cell growth and/or causes cell death, depending on basal class 3 ALDH activity. To confirm the correlation between inhibition of class 3 ALDH and reduction of cell proliferation, we exposed hepatoma cells to antisense oligonucleotides (ODNs) against ALDH3. In JM2 hepatoma cell line, with high ALDH3 activity, the exposure to antisense ODNs significantly decreases mRNA and enzyme activity (90%). At the same time, cell growth was reduced by about 70%. The results confirm that in hepatoma cells ALDH3 expression is closely related with cell growth, and that its inhibition is important in reducing the proliferation of hepatoma cells overexpressing ALDH3.     

Genome-wide identification and analysis of the aldehyde dehydrogenase (ALDH) gene superfamily of Gossypium raimondii

Background

Aldehyde dehydrogenases (ALDHs) are members of the NAD(P)⁺-dependent protein superfamily which catalyzes aliphatic and aromatic aldehyde oxidation to non-toxic carboxylic acids. ALDH genes may offer promise for improving plant adaptation to environmental stress. Recently, elucidated genome sequences of Gossypium raimondii provide a foundation for systematic identification and analysis of ALDH genes. To date, this has been accomplished for many plant species except G. raimondii.

Results

In this study, thirty unique ALDH sequences that code for 10 ALDH families were identified in the G. raimondii genome. Phylogenetic analysis revealed that ALDHs were split into six clades in G. raimondii, and ALDH proteins from the same families were clustered together. Phylogenetic relationships of ALDHs from 11 plant species suggest that ALDHs in G. raimondii shared the highest protein homology with ALDHs from poplar. Members within ALDH families possessed homologous exon–intron structures. Chromosomal distribution of ALDH did not occur evenly in the G. raimondii genome and many ALDH genes were involved in the syntenic region as documented by identification of physical locations among single chromosomes. In addition, syntenic analysis revealed that homologues of many G. raimondii ALDHs appeared in corresponding Arabidopsis and poplar syntenic blocks, indicating that these genes arose prior to G. raimondii, Arabidopsis and poplar speciation. Finally, based on gene expression analysis of microarray and RNA-seq, we can speculate that some G. raimondii ALDH genes might respond to drought or waterlogging stresses.

Conclusion

Genome-wide identification and analysis of the evolution and expression of ALDH genes in G. raimondii laid a foundation for studying this gene superfamily and offers new insights into the evolution history and speculated roles in Gossypium. These data can be used to inform functional genomic studies and molecular breeding in cotton.     

Molecular characterization of two alanine-rich Lea genes abundantly expressed in the resurrection plant C. plantagineum in response to osmotic stress and ABA

Expression of many genes is induced during dehydration in vegetative tissues of the desiccation tolerant resurrection plantCraterostigma plantagineum. The most abundant group of desiccation-related gene products belong to the LEA (= Late Embryogenesis Abundant) proteins. Here we describe structures and expression patterns of members of group 3 and group 4Lea genes fromC. plantagineum. The most intriguing observation is the strong conservation of repeat motifs inLea genes found across divers plant species includingC. plantagineum and non-desiccation tolerant plants. This conservation of structural elements leads to speculations about evolution of desiccation tolerance in the resurrection plant.     

Shifting the NAD/NADP preference in class 3 aldehyde dehydrogenase.

Among pyridine-nucleotide-dependent oxidoreductases, the class 3 family of aldehyde dehydrogenases (ALDHs) is unusual in its ability to function with either NAD or NADP. This is all the more surprising because an acidic residue, Glu140, coordinates the adenine ribose 2' hydroxyl. In many NAD-dependent dehydrogenases a similarly placed carboxylate is thought to be responsible for exclusion of NADP. The corresponding residue in most (approximately 71%) sequences in the ALDH extended family is also Glu, and most of these are NAD-specific enzymes. Site-directed mutagenesis was performed on this residue in rat class 3 ALDH. Our results indicate that this residue contributes to tighter binding of NAD in the native enzyme, but suggest that additional factors must contribute to the ability to utilize NADP. Mutagenesis of an adjacent basic residue (Lys137) indicates that it is even more essential for binding both coenzymes, consistent with its conservation in nearly all ALDHs (> 98%).     

Isoelectric focusing studies of aldehyde dehydrogenases, alcohol dehydrogenases and oxidases from mammalian anterior eye tissues

1. Isoelectric focusing (IEF) and zymogram methods were used to examine the tissue distribution, multiplicity and substrate specificities of alcohol dehydrogenases (ADHs), aldehyde dehydrogenases (ALDHs) and ocular oxidases (EOXs) from mammalian anterior eye tissues. 2. Baboon, cattle, pig and sheep corneal extracts exhibited high ALDH activities; the corneal ALDHs were distinct from the major liver ALDHs and distinguished by their preference for medium-chain aldehydes. 3. Baboon and pig corneal extracts also showed high ADH activities, by comparison with ovine and bovine samples. Moreover, the ADHs were distinct from the major liver isozymes in pI value and substrate specificity. 4. Mammalian lens extracts exhibited significant ALDH activity of a form corresponding to the major liver cytosolic isozyme. Minor activity of the corneal enzyme was also observed in some species. 5. Lens ADH phenotypes were species-specific, and consisted of either Class II activity (baboon and sheep), Class III ADH activity (pig), or activities of both ADH classes (cattle). 6. Lens extracts also exhibited a complex pattern of ocular oxidase (EOX) activities following IEF. 7. A role in peroxidatic aldehyde detoxification is proposed for these enzymes in anterior eye tissues.     

Apo and holo crystal structures of an NADP-dependent aldehyde dehydrogenase from Streptococcus mutans.

The aldehyde dehydrogenases (ALDHs) are a superfamily of multimeric enzymes which catalyse the oxidation of a broad range of aldehydes into their corresponding carboxylic acids with the reduction of their cofactor, NAD or NADP, into NADH or NADPH. At present, the only known structures concern NAD-dependent ALDHs. Three structures are available in the Protein Data Bank: two are tetrameric and the other is a dimer. We solved by molecular replacement the first structure of an NADP-dependent ALDH isolated from Streptococcus mutans, in its apo form and holo form in complex with NADP, at 1.8 and 2.6 A resolution, respectively. Although the protein sequence shares only approximately 30 % identity with the other solved tetrameric ALDHs, the structures are very similar. However, a large local conformational change in the region surrounding the 2' phosphate group of the adenosine moiety is observed when the enzyme binds NADP, in contrast to the NAD-dependent ALDHs.Structure and sequence analyses reveal several properties. A small number of residues seem to determine the oligomeric state. Likewise, the nature (charge and volume) of the residue at position 180 (Thr in ALDH from S. mutans) determines the cofactor specificity in comparison with the structures of NAD-dependent ALDHs. The presence of a hydrogen bond network around the cofactor not only allows it to bind to the enzyme but also directs the side-chains in a correct orientation for the catalytic reaction to take place. Moreover, a specific part of this network appears to be important in substrate binding. Since the enzyme oxidises the same substrate, glyceraldehyde-3-phosphate (G3P), as NAD-dependent phosphorylating glyceraldehyde-3-phosphate dehydrogenases (GAPDH), the active site of GAPDH was compared with that of the S. mutans ALDH. It was found that Arg103, Arg283 and Asp440 might be key residues for substrate binding.     

Characterization of the aldehyde dehydrogenase gene families of Zea mays and Arabidopsis

Cytoplasmic male sterility is a maternally transmitted inability to produce viable pollen. Male sterility occurs in Texas (T) cytoplasm maize as a consequence of the premature degeneration of the tapetal cell layer during microspore development. This sterility can be overcome by the combined action of two nuclear restorer genes, rf1 and rf2a. The rf2a gene encodes a mitochondrial aldehyde dehydrogenase (ALDH) that is capable of oxidizing a variety of aldehydes. Six additional ALDH genes were cloned from maize and Arabidopsis. In vivo complementation assays and in vitro enzyme analyses demonstrated that all six genes encode functional ALDHs. Some of these ALDHs are predicted to accumulate in the mitochondria, others in the cytosol. The intron/exon boundaries of these genes are highly conserved across maize and Arabidopsis and between mitochondrial and cytosolic ALDHs. Although animal, fungal, and plant genomes each encode both mitochondrial and cytosolic ALDHs, it appears that either the gene duplications that generated the mitochondrial and the cytosolic ALDHs occurred independently within each lineage or that homogenizing gene conversion-like events have occurred independently within each lineage. All studied plant genomes contain two confirmed or predicted mitochondrial ALDHs. It appears that these mitochondrial ALDH genes arose via independent duplications after the divergence of monocots and dicots or that independent gene conversion-like events have homogenized the mitochondrial ALDH genes in the monocot and dicot lineages. A computation approach was used to identify amino acid residues likely to be responsible for functional differences between mitochondrial and cytosolic ALDHs.     

Role of aldehyde dehydrogenases in endogenous and xenobiotic metabolism

Aldehydes are highly reactive molecules that are intermediates or products involved in a broad spectrum of physiologic, biologic and pharmacologic processes. Aldehydes are generated from chemically diverse endogenous and exogenous precursors and aldehyde-mediated effects vary from homeostatic and therapeutic to cytotoxic, and genotoxic. One of the most important pathways for aldehyde metabolism is their oxidation to carboxylic acids by aldehyde dehydrogenases (ALDHs). Oxidation of the carbonyl functional group is considered a general detoxification process in that polymorphisms of several human ALDHs are associated a disease phenotypes or pathophysiologies. However, a number of ALDH-mediated oxidation form products that are known to possess significant biologic, therapeutic and/or toxic activities. These include the retinoic acid, an important element for vertebrate development, gamma-aminobutyric acid (GABA), an important neurotransmitter, and trichloroacetic acid, a potential toxicant. This review summarizes the ALDHs with an emphasis on catalytic properties and xenobiotic substrates of these enzymes.     

Human aldehyde dehydrogenase-catalyzed oxidation of ethylene glycol ether aldehydes

Ethylene glycol ethers (EGEs) are primary alcohols commonly used as solvents in numerous household and industrial products. Exposure to EGEs has been correlated with delayed encephalopathy, metabolic acidosis, sub-fertility and spermatotoxicity in humans. In addition, they also cause teratogenesis, carcinogenesis, hemolysis, etc., in various animal models. Metabolism EGEs parallels ethanol metabolism, i.e., EGEs are first converted to 2-alkoxy acetaldehydes (EGE aldehydes) by alcohol dehydrogenases, and then to alkoxyacetic acids by aldehyde dehydrogenases (ALDHs). The acid metabolite of EGEs is considered responsible for toxicities associated with EGEs. The role of human ALDHs in EGE metabolism is not clear; accordingly, we have investigated the ability of five different human ALDHs (ALDH1A1, ALDH2, ALDH3A1, ALDH5A1 and ALDH9A1) to catalyze the oxidation of various EGE aldehydes. The EGE aldehydes used in this study were synthesized via Swern oxidation. All of the human ALDHs were purified from human cDNA clones over-expressing these enzymes in E. coli. The ALDHs tested, so far, differentially catalyze the oxidation of EGE aldehydes to their corresponding acids (K(m) values range from approximately 10 microM to approximately 20.0mM). As judged by V(max)/K(m) ratios, short-chain alkyl-group containing EGE aldehydes are oxidized to their acids more efficiently by ALDH2, whereas aryl- and long-chain alkyl-group containing EGE aldehydes are oxidized to their acid more efficiently by ALDH3A1. Given the product of ALDH-catalyzed reaction is toxic, this process should be considered as a bio-activation (toxification) process.     

Recombinant class I aldehyde dehydrogenases specific for all-trans- or 9-cis-retinal

The molecular basis for the specificity of aldehyde dehydrogenases (ALDHs) for retinal, the precursor of the morphogen retinoic acid, is still poorly understood. We have expressed in Escherichia coli both retinal dehydrogenase (RALDH), a cytosolic aldehyde dehydrogenase originally isolated from rat kidney, and the highly homologous phenobarbital-induced aldehyde dehydrogenase (PB-ALDH). Oxidation of propanal was observed with both enzymes. On the other hand, recombinant RALDH efficiently catalyzed oxidation of 9-cis- and all-trans-retinal, whereas PB-ALDH was inactive with all-trans-retinal and poorly active with 9-cis-retinal. A striking difference between PB-ALDH and all other class I ALDHs is the identity of the amino acid immediately preceding the active nucleophile Cys(302) (Ile(301) instead of Cys(301)). Nevertheless, these amino acids could be exchanged in either RALDH or PB-ALDH without affecting substrate specificity. Characterization of chimeric enzymes demonstrates that distinct groups of amino acids control the differential activity of RALDH and PB-ALDH with all-trans- and 9-cis-retinal. Of 52 divergent amino acids, the first 17 are crucial for activity with all-trans-retinal, whereas the next 25 are important for catalysis of 9-cis-retinal oxidation. Recombinant enzymes with specificity for all-trans- or 9-cis-retinal were obtained, which should provide useful tools to study the relative importance of local production of all-trans- versus 9-cis-retinoic acid in development and tissue differentiation.