CHARACTERIZATION OF AN ATP-DEPENDENT DNA-LIGASE ENCODED BY HAEMOPHILUS-INFLUENZAE

We report that Haemophilus influenzae encodes a 268 amino acid ATP-dep endent DNA ligase. The specificity of Haemophilus DNA ligase was inves tigated using recombinant protein produced in Escherichia coli. The en zyme catalyzed efficient strand joining on a singly nicked DNA in the presence of magnesium and ATP (K-m = 0.2 mu M) Other nucleoside tripho sphates or deoxynucleoside triphosphates could not substitute for ATP. Haemophilus ligase reacted with ATP in the absence of DNA substrate t o form a covalent ligase-adenylate intermediate. This nucleotidyl tran sferase reaction required a divalent cation and was specific for ATP. The Haemophilus enzyme is the first example of an ATP-dependent DNA li gase encoded by a eubacterial genome. It is also the smallest member o f the covalent nucleotidyl transferase superfamily, which includes the bacteriophage and eukaryotic ATP-dependent polynucleotide ligases and the GTP-dependent RNA capping enzymes.