Binding modes of inhibitors to ribonuclease T1 as elucidated by the analysis of two-dimensional NMR.
Aromatic proton and high field shifted methyl proton resonances of RNase T1 complexed with Guo, 2'GMP, 3'GMP or 5'GMP were assigned to specific amino acid residues by 2D-NMR spectra in comparison with the crystal structure of RNase T1-2'GMP complex. The spatial proximities of amino acid residues as elucidated by NOESY spectra were found to be quite similar among free RNase T1 and the inhibitor complexes, showing that large conformational changes did not occur upon complex formation. However, small but appreciable conformational changes were induced which were reflected by the systematic chemical shift changes of some amino acid residues in the active site. Furthermore, we confirmed that RNase T1 contains two specific binding sites, one for the guanine base and the other for the phosphate moiety. The inhibitors are forced to adapt their conformations to fit the guanine base and the phosphate moiety to each binding site on the enzyme. This is consistent with our previous studies that 2'GMP and 3'GMP take syn form as a bound conformation, while 5'GMP takes anti conformation around glycosidic bonds.
Shimada
I
I
Department of Molecular Physiology, Tokyo Metropolitan Institute of Medical Science, Japan.
Inagaki
F
F
D004722:Endoribonucleases / Q000037:antagonists & inhibitors*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D011485:Protein Binding; D006163:Ribonuclease T1 / Q000037:antagonists & inhibitors* / Q000378:metabolism
D004722:Endoribonucleases; D006163:Ribonuclease T1