A potential regulatory role of the N-terminal domain of Dna2 during Okazaki fragment maturation

초록

The Saccharomyces cerevisiae Dna2 protein involved in Okazaki fragment processing is a multifunctional enzyme. Deletion of its N-terminal 405 amino acids did not cause any loss of the two-enzymatic activities of Dna2; helicase and single-stranded DNA-specific endonuclease. However, cells with a mutant DNA2 allele (dna2D405N) lacking the N-terminal 405 amino acids were severely impaired in growth, indicating that the N-terminal region contains a domain critical for a cellular function(s) of Dna2. In order to understand the biological role of the N-terminal domain, we examined and compared the enzymological properties of Dna2D405N mutant enzyme with those of wild type one. Wild type Dna2 efficiently cleaved and unwound the substrates with trinucleotide repeated sequences (TNR), whereas, the mutant enzyme poorly utilized the substrates. Consistent with this, the purified N-terminal 405 amino acid protein bound to the substrate, but the mutant protein lacking this domain did not. We also observed instability of TNR in the dna2D405N mutant strain. This suggests Dna2 has a role in maintaining TNR in the eukaryotic cells, and the N-terminal domain is important for this function of Dna2.