Part:BBa_K5288003
Betaine aldehyde dehydrogenase
Description
The biobrick consists of a three-domain protein fused with a 6x His-tag to facilitate the purification process. Betaine aldehyde dehydrogenase (BADH) consists of three domains; an N-terminal NAD+ binding domain, an oligomerization domain, and a C-terminal catalytic domain [1], [2], [3]. The enzyme has a length of 510 amino acids and a molecular weight of 55.61 kDa. The part is adapted to the Golden Gate cloning method. This part also contains a x6 HisTag in the C-terminal site, to facilitate its purification process.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 198
Illegal PstI site found at 1635 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 198
Illegal NheI site found at 487
Illegal NheI site found at 736
Illegal PstI site found at 1635 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 198
- 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 198
Illegal PstI site found at 1635 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 198
Illegal PstI site found at 1635 - 1000COMPATIBLE WITH RFC[1000]
Usage and Biology
Hydric and heat stress occur when plants face limited water and high temperatures, severely impacting crop yields, sometimes reducing them by almost half, and often resulting in poor or unusable product. This leads to a shortage of key nutrients in basic foods. Current treatments, such as compost, humic acids, and hydrogels, are applied directly to the soil but can cause environmental issues or adverse effects if not used correctly [4], [5], [6]. Our team has been investigating a safe, efficient, and easy-to-use solution to boost crop resilience under these harsh conditions.
To address this problem, we designed an innovative solution that acts as an active and preventive method. A biostimulant based on the biosynthesis of three key osmolytes and osmoprotectants: Proline, Glycine Betaine, and Glucose. The production of these metabolites inside the plant will ensure an increase in water management.
The principle behind the biostimulant relies on the biosynthesis of osmolytes and osmoprotectants. Like electrolytes, osmolytes contribute to homeostasis maintenance. They provide the driving gradient for water uptake and maintain cell turgor by osmotic adjustment and redox metabolism to remove excess reactive oxygen species (ROS) and reestablish the cellular redox balance. Also, they protect cellular machinery from osmotic stress and oxidative damage [7]. Related to osmolytes, osmoprotectants can protect cells against osmotic stress and high salinity conditions. These compounds protect cells by stabilizing proteins, maintaining membrane integrity, and scavenging reactive oxygen species [8].
Glycine Betaine (GB) biosynthesis can happen via two substrates: choline oxidation or glycine methylation [9]. Upon higher plants, GB biosynthesis consists of a two-step, multi-enzymatic process [10], [11], [12], [13]. The first step is catalyzed by the Fe-dependent Choline monooxygenase (CMO), where choline is hydroxylated into Betaine aldehyde [9], [11], [12], [14]. Hydroxylation is irreversible [15]. Then Betaine aldehyde is oxidized by the NAD+-dependent Betaine aldehyde Dehydrogenase to produce GB [11], [12]. This process occurs within the chloroplasts [9], [10], [14], [16]. CMO plays a big role in the biosynthesis of GB, as is the rate-limiting enzyme [11], [14].
Betaine aldehyde dehydrogenases (BADHs) are common among life, they can be found in many plants, mammals, bacteria, and yeasts [1]. BADHs belong to the aldehyde dehydrogenases (ALDHs) superfamily. ALDHs are an evolutionary-conserved superfamily of oxidoreductases, which can convert a large array of aldehydes to carboxylic acids [17]. ALDHs have 24 families recognized in eukaryotes. ALDHs take a role in the oxidation of amino aldehydes [11], [18]. BADH for animals, fungi, and proteobacteria is classified into the ALDH9 family. In contrast, functionally BADHs from plants are classified as ALDH10 family [3], [13], [19]. Even though there is an evolutionary separation between these two families, ALDH9 and ALDH10 have a 39% similarity [18]. Also, plants produced BADH-like aldehyde reductases classified in the ALDH22 family [13]. Moreover, plants have two isogenes for BADH, with the hypothesized difference in its localization and functionality [1], [20], [21]. In plants, BADH can catalyze the oxidation of other aldehydes [22]. Also, BADH plays a role in the production of fragrance compounds and detoxification [18], [23].
Plants BADHs are mainly composed of three well-conserved domains, an N-terminal NAD+ binding domain, an oligomerization domain, and a C-terminal catalytic domain [1], [2], [3]. Specifically, this biobrick is the coding sequence for BADH from Zea mays (LOC541949).
The N-terminal NAD+ binding domain is common among ALDHs [17]. This domain has a Rossman fold structure. Rossman folds are ancient and structurally diverse folds present in a wide range of proteins. It belongs to the doubly wound superfold, which is one of the most prevalent superfolds in nature [24], [25]. Rossman folds are composed basically of a series of alternating beta strands (β) and alpha-helical (α) structures. Rossman folds domains are likely to bind to the ADP portion of dinucleotides like FAD, NAD, and NADP [25]. Especially the N-terminal domain has a preference for NAD+, as in most of the ALDHs [17].
The C-terminal substrate binding or catalytic domain is rich in cysteines, which gives the affinity to the substrate. This same characteristic can be targeted by the drug Disulfiram, which oxidizes the cysteines and inhibits the enzyme [19].
We intend to use the BADH as a key enzyme for the biosynthesis of the osmolyte and osmoprotectant; GB. As mentioned previously, CMO catalyzes the second step in the biosynthesis of GB. By this, we intend to ensure a better resistance to drought and caloric stress on corn. GB is an environmentally safe, non-toxic soluble compound [26]. GB stabilizes membranes and protects proteins while being under dehydration stress [10]. Furthermore, GB improves osmotic adjustment and stabilizes subcellular structures, preserving the thermodynamic stability of macromolecules and reversing protein misfolding without compromising native protein functionality. The high solubility and hydrophilicity of GB aid in maintaining the water contents of plants as much as possible in arid environments [27]. Even more, GB stimulates the production of ROS-scavenging enzymes. GB accumulation, in maize, is demonstrated to enhance the germination process and increase the number of antioxidant-related enzymes [28]. This will lead to a novel non-transgenic or synthetic solution to losses on corn production globally due to abiotic stress.
References
[1] M. Chen et al., “CRISPR/Cas9-Mediated Targeted Mutagenesis of Betaine Aldehyde Dehydrogenase 2 (BADH2) in Tobacco Affects 2-Acetyl-1-pyrroline”, Agronomy, vol. 14, núm. 2, p. 321, feb. 2024, doi: 10.3390/agronomy14020321.
[2] N. A. Stephens-Camacho, A. Muhlia-Almazan, A. Sanchez-Paz, y J. A. Rosas-Rodríguez, “Surviving environmental stress: the role of betaine aldehyde dehydrogenase in marine crustaceans”, Invertebr. Surviv. J., vol. 12, núm. 1, Art. núm. 1, feb. 2015.
[3] C. Muñoz‐Bacasehua et al., “Heterogeneity of active sites in recombinant betaine aldehyde dehydrogenase is modulated by potassium”, J. Mol. Recognit., vol. 33, núm. 10, p. e2869, oct. 2020, doi: 10.1002/jmr.2869.
[4] R. F. Ratke et al., “Cashew gum hydrogel as an alternative to minimize the effect of drought stress on soybean”, Sci. Rep., vol. 14, núm. 1, p. 2159, ene. 2024, doi: 10.1038/s41598-024-52509-2.
[5] R. Hernández-Campos, C. Robles, y A. Calderín García, “EFECTO DE ÁCIDOS HÚMICOS EN EL CRECIMIENTO VEGETAL Y LA PROTECCIÓN CONTRA EL ESTRÉS HÍDRICO EN POBLACIONES SELECCIONADAS DE MAÍZ NATIVO DE MÉXICO”, Rev. Fitotec. Mex., vol. 44, núm. 4, p. 561, dic. 2021, doi: 10.35196/rfm.2021.4.561.
[6] F. Hellal, S. El Sayed, D. M. R. A. Basha, y H. H. A. Kader, “Mitigation of water stress by compost and arginine application and its impacts on barley production”, Bull. Natl. Res. Cent., vol. 48, núm. 1, p. 25, feb. 2024, doi: 10.1186/s42269-024-01178-2.
[7] U. K. Ghosh, Md. N. Islam, Md. N. Siddiqui, y Md. A. R. Khan, “Understanding the roles of osmolytes for acclimatizing plants to changing environment: a review of potential mechanism”, Plant Signal. Behav., vol. 16, núm. 8, p. 1913306, ago. 2021, doi: 10.1080/15592324.2021.1913306.
[8] D. Mehta y S. Vyas, “Comparative bio-accumulation of osmoprotectants in saline stress tolerating plants: A review”, Plant Stress, vol. 9, p. 100177, sep. 2023, doi: 10.1016/j.stress.2023.100177.
[9] F. Zulfiqar, M. Ashraf, y K. H. M. Siddique, “Role of Glycine Betaine in the Thermotolerance of Plants”, Agronomy, vol. 12, núm. 2, p. 276, ene. 2022, doi: 10.3390/agronomy12020276.
[10] P. Suraninpong, K. Thongkhao, A. M. Azzeme, y P. Suksa-Ard, “Monitoring Drought Tolerance in Oil Palm: Choline Monooxygenase as a Novel Molecular Marker”, Plants, vol. 12, núm. 17, p. 3089, ago. 2023, doi: 10.3390/plants12173089.
[11] Z. Xu et al., “Glycinebetaine Biosynthesis in Response to Osmotic Stress Depends on Jasmonate Signaling in Watermelon Suspension Cells”, Front. Plant Sci., vol. 9, p. 1469, oct. 2018, doi: 10.3389/fpls.2018.01469.
[12] S. S. Krishnamurthi, S. Kuttan, T. Nooruddin, y A. Parida, “The Role of the Overexpression of Suaeda maritima Choline Monooxygenase and Betaine Aldehyde Dehydrogenase cDNAs in the Enhancement of Salinity Tolerance in Different Strains of”, vol. 12, núm. 1, 2019.
[13] R. Ming, Y. Zhang, Y. Wang, M. Khan, B. Dahro, y J. Liu, “The JA‐responsive MYC2‐ BADH ‐ like transcriptional regulatory module in Poncirus trifoliata contributes to cold tolerance by modulation of glycine betaine biosynthesis”, New Phytol., vol. 229, núm. 5, pp. 2730–2750, mar. 2021, doi: 10.1111/nph.17063.
[14] Y. Wang et al., “Leveraging Atriplex hortensis choline monooxygenase to improve chilling tolerance in cotton”, Environ. Exp. Bot., vol. 162, pp. 364–373, jun. 2019, doi: 10.1016/j.envexpbot.2019.03.012.
[15] J. Carrillo-Campos, H. Riveros-Rosas, R. Rodríguez-Sotres, y R. A. Muñoz-Clares, “Bona fide choline monoxygenases evolved in Amaranthaceae plants from oxygenases of unknown function: Evidence from phylogenetics, homology modeling and docking studies”, PLOS ONE, vol. 13, núm. 9, p. e0204711, sep. 2018, doi: 10.1371/journal.pone.0204711.
[16] E. M. Valenzuela-Soto y C. G. Figueroa-Soto, “Biosynthesis and Degradation of Glycine Betaine and Its Potential to Control Plant Growth and Development”, en Osmoprotectant-Mediated Abiotic Stress Tolerance in Plants, M. A. Hossain, V. Kumar, D. J. Burritt, M. Fujita, y P. S. A. Mäkelä, Eds., Cham: Springer International Publishing, 2019, pp. 123–140. doi: 10.1007/978-3-030-27423-8_5.
[17] N. Stiti, V. Giarola, y D. Bartels, “From algae to vascular plants: The multistep evolutionary trajectory of the ALDH superfamily towards functional promiscuity and the emergence of structural characteristics”, Environ. Exp. Bot., vol. 185, p. 104376, may 2021, doi: 10.1016/j.envexpbot.2021.104376.
[18] F. Jacques et al., “Roles for ALDH10 enzymes in γ-butyrobetaine synthesis, seed development, germination, and salt tolerance in Arabidopsis”, J. Exp. Bot., vol. 71, núm. 22, pp. 7088–7102, dic. 2020, doi: 10.1093/jxb/eraa394.
[19] R. Cruz-Valencia, A. A. Arvizu-Flores, J. A. Rosas-Rodríguez, y E. M. Valenzuela-Soto, “Effect of the drug cyclophosphamide on the activity of porcine kidney betaine aldehyde dehydrogenase”, Mol. Cell. Biochem., vol. 476, núm. 3, pp. 1467–1475, mar. 2021, doi: 10.1007/s11010-020-04010-3.
[20] T. Fujiwara et al., “Enzymatic characterization of peroxisomal and cytosolic betaine aldehyde dehydrogenases in barley”, Physiol. Plant., vol. 134, núm. 1, pp. 22–30, sep. 2008, doi: 10.1111/j.1399-3054.2008.01122.x.
[21] D. Luo et al., “Rice choline monooxygenase (OsCMO) protein functions in enhancing glycine betaine biosynthesis in transgenic tobacco but does not accumulate in rice (Oryza sativa L. ssp. japonica)”, Plant Cell Rep., vol. 31, núm. 9, pp. 1625–1635, sep. 2012, doi: 10.1007/s00299-012-1276-2.
[22] T. Nakamura, M. Nomura, H. Mori, A. T. Jagendorf, A. Ueda, y T. Takabe, “An Isozyme of Betaine Aldehyde Dehydrogenase inBarley”, Plant Cell Physiol., vol. 42, núm. 10, pp. 1088–1092, oct. 2001, doi: 10.1093/pcp/pce136.
[23] A. Maghraby y M. Alzalaty, “Genome-wide identification, characterization and evolutionary analysis of betaine aldehyde dehydrogenase (BADH), mitogen-activated protein kinase (MAPK) and sodium/hydrogen exchanger (NHX) genes in maize (Zea mays) under salt stress”, Genet. Resour. Crop Evol., mar. 2024, doi: 10.1007/s10722-024-01930-7.
[24] K. E. Medvedev, L. N. Kinch, R. Dustin Schaeffer, J. Pei, y N. V. Grishin, “A Fifth of the Protein World: Rossmann-like Proteins as an Evolutionarily Successful Structural unit”, J. Mol. Biol., vol. 433, núm. 4, p. 166788, feb. 2021, doi: 10.1016/j.jmb.2020.166788.
[25] I. Hanukoglu, “Proteopedia: Rossmann fold: A beta‐alpha‐beta fold at dinucleotide binding sites”, Biochem. Mol. Biol. Educ., vol. 43, núm. 3, pp. 206–209, may 2015, doi: 10.1002/bmb.20849.
[26] S. Ali et al., “Glycine Betaine Accumulation, Significance and Interests for Heavy Metal Tolerance in Plants”, Plants, vol. 9, núm. 7, p. 896, jul. 2020, doi: 10.3390/plants9070896.
[27] M. Bai et al., “Transcriptome expression profiles reveal response mechanisms to drought and drought-stress mitigation mechanisms by exogenous glycine betaine in maize”, Biotechnol. Lett., vol. 44, núm. 3, pp. 367–386, mar. 2022, doi: 10.1007/s10529-022-03221-6.
[28] M. Niazian, S. A. Sadat-Noori, M. Tohidfar, S. M. M. Mortazavian, y P. Sabbatini, “Betaine Aldehyde Dehydrogenase (BADH) vs. Flavodoxin (Fld): Two Important Genes for Enhancing Plants Stress Tolerance and Productivity”, Front. Plant Sci., vol. 12, p. 650215, abr. 2021, doi: 10.3389/fpls.2021.650215.
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