Snapshots of Motion: A Novel Structural Intermediate Reveals Conserved Dynamics in Archaeal DNA Ligases

We present the first structural evidence of an archaeal ligase in complex with AMP, displaying the covalent bond between the ε-amino group of the enzyme and the α-phosphate of the AMP. This interaction captures a structural intermediary in the first step of the ligation reaction, substantiating further cofactor hydrolysis as a trigger for domain rearrangement leading to the formation of the catalytic cavity. Our crystallographic models of Thermococcus gammatolerans DNA ligase (LigTgam), coupled with bioinformatic analysis of at least 28 crystallographic structures from ATP- and NAD+-dependent DNA ligases, highlight the central role of domain mobility. Notably, elevated B-values are consistently observed in key catalytic and binding regions, suggesting a link between structural flexibility and enzymatic efficiency. Remarkably, this pattern of high B-values is conserved in replicative ligases, including bacterial Lig A, indicating a broader evolutionary relevance. These fluctuations emphasize the importance of conformational adaptability in accommodating substrate DNA and facilitating catalytic steps, including adenylation and phosphodiester bond formation. In this work, we delve deeper into this dynamic behavior, providing evidence of its critical role in ligase function.