NMR Studies on N-terminal Domain of DNA2

Abstract

Saccharomyces cerevisiae Dna2 protein has biochemical activities: DNA-dependent ATPase, DNA helicase and DNA nuclease and is essential for cell viability. Especially, Pro504 is determined as an important residue in ATPase, helicase, and nuclease activity. We synthesized and determined the three-dimensional solution structure of N-terminal domain comprising residues of Val501-Phe508 (Dna2pep) using two-dimensional 1H-NMR and dynamical simulated annealing calculations. On the basis of a total of 44 experimental restraints including NOEs, 3Jand 3J coupling constants, the solution structures of Dna2pep were calculated with the program CNS. The 23 lowest energy structures were selected out of 50 final simulated-annealing structures. The atomic RMSDs of the final 23 structures for the individual residues were calculated with respect to the average structure. The mean RMSDs for the 23 structures were 0.042 nm for backbone atoms and 0.316 nm for all heavy atoms, respectively. The Ramachandran plot indicates that the ,  angles of the 23 final structures are properly distributed in energetically acceptable regions. Solution structure of Dna2pep showed a single unique turn spanning residues of Asn503 – Val506.