A Cold-Induced LEA3 Protein, DohD, Confers Cryoprotective Protection Against Low-Temperature Stress in .

is a remarkably unique microorganism, exhibiting extraordinary tolerance to extreme conditions such as ionizing radiation, ultraviolet light, and desiccation. However, the response mechanisms of under low-temperature stress remain largely unexplored and have yet to be fully elucidated. The DohD protein is a hydrophilic member of the late embryogenesis abundant 3 (LEA3) family of , playing a pivotal role in abiotic stress adaptation. Bioinformatics analysis revealed that DohD contains tandem repeats and disordered domains, with a remarkably high α-helix content (91.41%). Furthermore, DohD exhibits extremely low homology with other proteins, highlighting its uniqueness to . Under low-temperature stress (15 °C), the expression of was significantly upregulated (5-fold), regulated by a dual mechanism involving positive control by DrRRA and negative regulation by Csp. Circular dichroism spectroscopy unveiled temperature-dependent structural plasticity: as the temperature increased from 0 °C to 50° C, the α-helix content decreased from 23.5% to 18.7%, while the antiparallel β-sheet content increased from 31.3% to 50.8%. This suggests an α-helix to β-sheet interconversion mechanism as a strategy for thermal adaptation. Additionally, deletion of impaired the tolerance of to cold, desiccation, oxidative, and high-salt stresses, accompanied by the reduced activities of antioxidant enzymes (SOD, CAT, POD) and the downregulation of related gene expression. This study elucidates the multifunctional role of DohD in stress resistance through structural dynamics, transcriptional regulation, and redox homeostasis, providing valuable insights into the adaptation mechanisms of extremophiles.