Nuclear magnetic resonance study of the interaction of T4 endonuclease V with DNA.
T4 endonuclease V catalyzes the DNA strand cleavage in the vicinity of a thymine dimer. In order to obtain insight into the specific recognition mechanism of this enzyme with a thymine photodimer within DNA, the conformations of five different DNA duplexes, [sequence: see text] with which the enzyme can interact, were studied by 1H NMR. DNA I, DNA IV, and DNA V do not contain the TT sequence or a thymine dimer and hence, are expected to bind the enzyme only in a nonspecific manner. DNA II includes a single TT sequence which does not form a thymine dimer. Only DNA III is expected to bind specifically to the enzyme through a thymine photodimer. The NMR spectra of these five DNA duplexes in the absence of the enzyme clearly show that the formation of a thymine dimer within the DNA induces only a minor distortion in the structure and that the overall structure of B-type DNA is retained. The photodimer formation is found to cause a large change in chemical shifts at the GC7 base pair, which is located at the 3'-side of the thymine dimer, accompanied by the major conformational change at the thymine dimer site. The effects of T4 endonuclease V binding on these DNA duplexes were also investigated by 1H NMR. The binding of this enzyme to DNA I, DNA IV, and DNA V causes no alteration in chemical shift values of the imino proton resonances, but the binding to DNA II induces a small downfield shift in the imino proton resonance of GC7.(ABSTRACT TRUNCATED AT 250 WORDS)
Lee
B J
BJ
Protein Engineering Research Institute, Osaka, Japan.
Sakashita
H
H
Ohkubo
T
T
Ikehara
M
M
Doi
T
T
Morikawa
K
K
Kyogoku
Y
Y
Osafune
T
T
Iwai
S
S
Ohtsuka
E
E
D001482:Base Composition; D001483:Base Sequence; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004706:Endodeoxyribonucleases / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011740:Pyrimidine Dimers; D013329:Structure-Activity Relationship; D014764:Viral Proteins
D009838:Oligodeoxyribonucleotides; D011740:Pyrimidine Dimers; D014764:Viral Proteins; D004247:DNA; D004706:Endodeoxyribonucleases; C470698:endonuclease V, phage T4; D043224:Deoxyribonuclease (Pyrimidine Dimer)