The discovery of DNA ligases in 1967 marked a crucial moment in molecular biology, with their role in joining 3'-OH and 5'-PO4 termini to form phosphodiester bonds making them indispensable for genome integrity. DNA ligases play vital roles in DNA replication and repair across all organisms. In recent years, research on DNA ligases has witnessed a resurgence, uncovering their diverse functions in various DNA repair pathways and their association with human genetic diseases. This article explores the intricate mechanisms of DNA ligases, their structural diversity, and the potential therapeutic applications arising from a deeper understanding of their functions.
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DNA Ligation Mechanism
The ligation of DNA involves a three-step nucleotidyl transfer process. Initially, ligase reacts with ATP or NAD+ to form a covalent ligase-adenylate intermediate. Subsequently, this intermediate transfers the AMP to the 5'-end of the 5'-phosphate-terminated DNA strand, creating DNA-adenylate. In the final step, ligase catalyzes the attack by the 3'-OH of the nick on DNA-adenylate, sealing the polynucleotides and releasing AMP. This intricate pathway of DNA sealing relies on divalent cation cofactors.
Fig. 1 Three-step pathway of nick sealing by DNA ligase (Shuman S. 2009).
Structural Diversity of DNA Ligases
DNA ligases can be classified into two families based on the substrate required for ligase-adenylate formation: ATP-dependent ligases and NAD+-dependent ligases. The former, represented by Chlorella virus DNA ligase (ChVLig), features an N-terminal nucleotidyltransferase (NTase) domain and a C-terminal OB domain. ChVLig serves as a minimalistic yet efficient ligase, forming a C-shaped protein clamp around the nicked duplex. Structural analysis reveals a significant domain rearrangement upon DNA binding, emphasizing the dynamic nature of ligase function.
In contrast, NAD+-dependent ligases, exemplified by Escherichia coli LigA, display a modular architecture with additional domains like Ia, a tetracycline zinc finger, an HhH domain, and a BRCT domain. LigA forms a unique clamp structure by engaging in kissing contacts between the NTase and HhH domains. The distinct structural features of ATP-dependent and NAD+-dependent ligases highlight the evolutionary divergence in their mechanisms of DNA envelopment.
Targeting DNA Ligases for Therapeutic Intervention
The essential role of DNA ligases in bacterial viability makes them attractive targets for antibacterial drug development. NAD+-dependent ligases, such as E. coli LigA, exhibit a druggable active site, providing an opportunity for the design of small molecule inhibitors. Recent studies have identified promising inhibitors that act competitively with NAD+, paving the way for potential antibacterial therapies.
Similarly, human DNA ligases present opportunities for therapeutic targeting in cancer treatment. Inhibiting DNA repair pathways by transiently impeding DNA ligase activity could sensitize tumor cells to DNA-damaging agents, potentially allowing for lower doses of chemotherapy and minimizing off-target effects.
In summary, DNA ligases, as guardians of genomic integrity, play pivotal roles in maintaining cellular functions. The diverse structural and biochemical characteristics of ligases offer insights into their evolutionary divergence and potential therapeutic applications. The ongoing research in this field holds the promise of uncovering new facets of DNA repair mechanisms, facilitating the development of targeted therapies for bacterial infections and cancer treatment. As our understanding deepens, the prospect of engineering designer ligases may open new frontiers in synthetic biology and genome editing.
Reference
- Shuman S. DNA ligases: progress and prospects. Journal of Biological Chemistry. 2009, 284(26): 17365-17369.
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