Secondary structure characterization of thermal adaptation in cytosolic malate dehydrogenase (cMDH) from three...

Understanding the thermal adaptation of protein function is key to predicting species distributions under climate warming. This study investigated the secondary structure of cytosolic malate dehydrogenase (cMDH) from three intertidal snails with distinct heat tolerance. Despite minimal sequence differences (2-3 amino acids) in cMDHs, their heat tolerance differed significantly. Circular dichroism spectra of wild-type and experimentally-generated mutant cMDHs confirmed that all are α-helix-rich proteins. The cMDH from the extremely heat-tolerant Echinolittorina malaccana had a significantly higher α-helix content than that from heat-tolerant E. radiata. A substitution from glycine to serine at residue 48 significantly reduced the α-helix content of cMDH (p < 0.05), however, substitutions at residues 4 and 114 did not significantly alter α-helix content (p > 0.05). Furthermore, the extremely heat-tolerant cMDH exhibited fewer salt bridges and hydrogen bonds, suggesting greater structural flexibility that may facilitate functional dynamics under heat stress. The findings demonstrate that the secondary structure content, particularly α-helix percentage, may be associated with the temperature adaptation of cMDHs.