Aldehyde dehydrogenases (ALDHs) comprise a protein superfamily of NAD(P)+-dependent enzymes (EC 1.2.1.-) and catalyze irreversible oxidation of aldehydes to carboxylic acids. ALDHs play a crucial role in detoxifying aldehydes produced by various metabolic pathways and during various stress conditions such as salinity, heat, cold and drought. At least 13 distinct families have been found in plants. Genomes of primitive plants such as algae comprise often a single ALDH gene per family while those of higher plants contain one to several ALDH gene copies per family. Gene duplication is a predominant evolutionary force behind existence of new genes, which undergo either partitioning of gene function or development of a new function. As plant organisms moved to land, there was a need for gene expansion linked to adaptation to new terrestrial stressors as well as to vascularization, tissue distribution and differentiation or regulation of hormone responses. As many studies on plants revealed that up-regulation of ALDH genes is a common target of stress response pathways, there is an economical interest in utilizing ALDH expression to improve crop growth and quality.
Here, we focused on ALDH10 and ALDH12 family members from moss P. patens, A. thaliana and maize (Z. mays) as well as ALDH22 family from maize and barley (H. vulgare) and ALDH23 from moss. Members of plant ALDH10 family are known to oxidize various aminoaldehydes arising from polyamine oxidation while ALDH12 oxidize semialdehyde dehydrogenase arising upon proline oxidation. So far, metabolites linked to ALDH22 and ALDH23 are unknown. The optimal buffer composition was analyzed using a nano differential scanning fluorimetry to achieve the highest stability of particular ALDH isoform. Substrate specificity was further evaluated spectrophotometrically, the best substrates were identified and kinetic constants were determined. Affinities towards the best aldehyde substrates and coenzyme NAD+ were further determined using microscale thermophoresis. The crystal structure of ZmALDH12 was solved at 2.2 Å resolution and subsequent site-directed mutagenesis was performed to identify key residues affecting the substrate and coenzyme preferences.
Supported by the grant 18-07563S from the Czech Science Foundation, young researcher grant JG_2020_001 from Palacký University, ERDF grant project No. CZ.02.1.01/0.0/0.0/16_019/0000827 (Plants as a tool for sustainable global development) from the Ministry of Education, Youth and Sports, Czech Republic.