DNA ligase

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ligase I, DNA, ATP-dependent
DNA ligase repairing chromosomal damage.
Identifiers
SymbolLIG1
Entrez3978
HUGO6598
OMIM126391
RefSeqNM_000234
UniProtP18858
Other data
EC number6.5.1.1
LocusChr. 19 [1]
ligase III, DNA, ATP-dependent
Identifiers
SymbolLIG3
Entrez3980
HUGO6600
OMIM600940
RefSeqNM_002311
UniProtP49916
Other data
LocusChr. 17 q11.2-q12
ligase IV, DNA, ATP-dependent
Identifiers
SymbolLIG4
Entrez3981
HUGO6601
OMIM601837
RefSeqNM_002312
UniProtP49917
Other data
LocusChr. 13 q33-q34

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Overview

In molecular biology, DNA ligase is a particular type of ligase (EC 6.5.1.1) that can link together DNA strands that have double-strand breaks (a break in both complementary strands of DNA). The alternative, a single-strand break, is easily fixed by DNA polymerase using the complementary strand as a template but still requires DNA ligase to create the final phosphodiester bond to fully repair the DNA.

DNA ligase has applications in both DNA repair and DNA replication (see Mammalian ligases). In addition, DNA ligase has extensive use in molecular biology laboratories for Genetic recombination experiments (see Applications in molecular biology research).

Ligase mechanism

The mechanism of DNA ligase is to form covalent phosphodiester bonds between 3' hydroxyl ends of one nucleotide with the 5' phosphate end of another. ATP is required for the ligase reaction.

A pictorial example of how a ligase works (with sticky ends): Error creating thumbnail: File missing

Ligase will also work with blunt ends, although higher enzyme concentrations and different reaction conditions are required.

Mammalian ligases

In mammals, there are four specific types of ligase.

Applications in molecular biology research

DNA ligases have become an indispensable tool in modern molecular biology research for generating recombinant DNA sequences. For example, DNA ligases are used with restriction enzymes to insert DNA fragments, often genes, into plasmids.

One vital, and often tricky, aspect to performing successful recombination experiments involving ligase is controlling the optimal temperature. Most experiments use T4 DNA Ligase (isolated from bacteriophage T4) which is most active at 25°c. However in order to perform successful ligations, the optimal enzyme temperature needs to be balanced with the melting temperature Tm (also the annealing temperature) of the DNA fragments being ligated.

If the ambient temperature exceeds Tm, homologous pairing of the sticky ends will not occur because the high temperature disrupts hydrogen bonding. The shorter the DNA fragments, the lower the Tm. Thus for sticky ends (overlaps) less than ten base pairs long, ligation experiments are performed at very low temperatures (~4-8°c) for a long period of time (often overnight).

The common commercially available DNA ligases were originally discovered in bacteriophage T4, E. coli or other bacteria.

See also

External links

da:Ligase de:Ligasen nl:DNA-ligase

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