Abstract
Aldehyde dehydrogenase (ALDH) is a widely recognized oxidoreductase that converts toxic aldehydes into harmless carboxylic acids, making it highly valuable for industrial applications. However, the effectiveness of ALDHs derived from Rhodococcus species in processing a range of aliphatic and aromatic aldehydes is still largely unexamined. Therefore, we cloned the ALDH gene from the cold-adapted strain Rhodococcus sp. PAMC28705 to address this gap and subsequently identified the crystal structure of rhALDH. By analyzing the unique structural features of the rhALDH active site, we evaluated its ability to process a wide range of aldehydes, with a focus on substrate specificity. Biochemical characterization revealed that at an optimal temperature of 30°C and a pH of 8.0, it exhibited the highest catalytic efficiency, with a kcat/Km value of 1.12 μM⁻¹ s⁻¹ for propionaldehyde, which was higher than that of its homologous ALDHs. This indicates a strong affinity for this substrate, as demonstrated by a low Km of 321.9 μM and a rapid turnover rate kcat of 359.2 s⁻¹. Adding disulfide reductants, such as dithiothreitol, 2-mercaptoethanol, and the metal ion Mg2+, further enhanced its activity. Working at mesophilic temperatures with good stability and substrate-specific catalytic efficiency, this novel rhALDH, which favors the conversion of propionaldehyde and benzaldehyde, provides a promising catalyst for biotechnological and sustainable bio-aldehyde elimination technologies. Thus, this study lays a foundation for future structure-function analyses for the development of ALDH-targeted antibiotics and the conservation of the environment from aldehyde toxicity.
