Interaction of T4 endonuclease V with DNA. Importance of the flexible loop regions in protein-DNA interaction

Hee Chul Ahn, Tadayasu Ohkubo, Shigenori Iwai, Kosuke Morikawa, Bong Jin Lee

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

T4 endonuclease V (T4 endo V), a thymine dimer-specific DNA repair enzyme, and its interaction with DNA were investigated by nuclear magnetic resonance (NMR) spectroscopy. Backbone resonance assignment, chemical shift mapping, and 15N relaxation measurements were employed to the free and DNA-bound enzymes. The secondary structure and the tertiary fold of T4 endo V in solution were consistent with those from the crystallographic study. The backbone 1H and 15N chemical shift perturbation upon the addition of DNA without a lesion revealed that the residues including Arg3, Arg22-Arg26, Lys45-Phe60, and Lys86-Thr88 participate in DNA binding. However, when DNA with a lesion was added to the enzyme and concomitantly the catalytic reaction was completed, the resonances of Arg22, Glu23, and Arg26, which constitute the catalytic active site, and the resonance of Thr88, were perturbed in a different manner. The region around Lys45-Ser47 was found to be involved in DNA binding, which have not been reported elsewhere. The backbone relaxation measurements of the free and DNA-bound enzymes indicated that two loop regions, Lys 45-Phe60 and Lyss86-Asp92, show the high degree of backbone flexibility. These results imply that two flexible loop regions may play an important role in DNA binding and in scanning along DNA duplex to search the thymine dimer sites in UV-damaged DNA.

Original languageEnglish
Pages (from-to)30985-30992
Number of pages8
JournalJournal of Biological Chemistry
Volume278
Issue number33
DOIs
StatePublished - 15 Aug 2003

Fingerprint

Dive into the research topics of 'Interaction of T4 endonuclease V with DNA. Importance of the flexible loop regions in protein-DNA interaction'. Together they form a unique fingerprint.

Cite this