- Escherichia coli (E. coli) DNA ligase is an essential enzyme in Escherichia coli that catalyzes the formation of phosphodiester bonds between adjacent nucleotides in DNA, thereby sealing nicks in the sugar-phosphate backbone during processes such as DNA replication, recombination, and repair. Unlike T4 DNA ligase, which uses ATP as a cofactor, E. coli DNA ligase specifically uses NAD⁺ (nicotinamide adenine dinucleotide) to activate the ligation reaction, making it distinct among known DNA ligases.
- This ligase catalyzes the joining of a 5′-phosphate and a 3′-hydroxyl group in double-stranded DNA substrates, but it exhibits high efficiency primarily on cohesive (sticky) ends and single-stranded nicks in DNA rather than blunt-ended fragments. The reaction mechanism involves the formation of a covalent enzyme–adenylate intermediate (ligase–AMP), transfer of the AMP to the 5′-phosphate of the DNA, and subsequent attack by the 3′-OH to form the phosphodiester bond, releasing AMP.
- E. coli DNA ligase plays a vital role in maintaining genomic integrity during cell growth and division. In molecular biology, it is less commonly used than T4 DNA ligase due to its limited activity on blunt-ended DNA and its NAD⁺ dependence, which may be less convenient in vitro. However, it remains valuable in specific applications requiring NAD⁺-dependent ligation systems, or in reconstitution of native E. coli DNA processing machinery for biochemical studies.
- Structurally, the enzyme is composed of conserved domains responsible for nucleotide binding and catalysis, similar to other ligases, and it shares evolutionary homology with NAD⁺-dependent ligases found in other bacteria. Research into E. coli DNA ligase and its mechanisms has provided foundational insight into ligation chemistry and has aided in the development of engineered ligases with modified substrate specificities.
- In summary, E. coli DNA ligase is a key bacterial enzyme specialized for NAD⁺-dependent DNA ligation during DNA metabolism. While not as broadly used in genetic engineering as T4 DNA ligase, it remains important in both bacterial physiology and biochemical research.
