Observation of arginyl-deoxyoligonucleotide interactions in Taq I endonuclease by detection of specific 1H NMR signals from 140kD [N eta 1, N eta 2, 15N Arg]Taq I/oligomer complexes.
Proton and nitrogen signals of the guanidinium amines in [N eta 1, N eta 2 15N Arg]Taq I endonuclease were observed using isotope filtered experiments and proton detected 1H[15N] heterocorrelated two dimensional NMR spectroscopy. These rapidly exchanging protons could be detected in the free enzyme only at pH 4.5; at pH 8.5, no signals were measured after extensive signal averaging. Addition of deoxyribonucleotide oligomers resulted in the appearance of two groups of signals at about 6.8 and 7.5 ppm. Since these signals are independent of the presence of cognate sequence or Mg2+, it is assumed they represent nonspecific arginyl-DNA interactions. This labeling/NMR approach provides a new method for investigating the role of arginine in protein-DNA interactions.
Glushka
J
J
Rockefeller University, N.Y., N.Y., 10021.
Barany
F
F
Cowburn
D
D
D001120:Arginine / Q000032:analysis*; D004247:DNA / Q000032:analysis*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000032:analysis*; D004926:Escherichia coli / Q000201:enzymology; D009682:Magnetic Resonance Spectroscopy; D009838:Oligodeoxyribonucleotides / Q000032:analysis*
D009838:Oligodeoxyribonucleotides; D004247:DNA; D001120:Arginine; D015252:Deoxyribonucleases, Type II Site-Specific; C027046:TCGA-specific type II deoxyribonucleases