Nature of the chemical bond formed with the structural metal ion at the A9/G10.1 motif derived from hammerhead ribozymes.
We have studied the interaction between metal ions and the metal ion-binding motif in hammerhead ribozymes, as well as the functions of the metal ion at the motif, with heteronuclear NMR spectroscopy. In this study, we employed model RNA systems which mimic the metal ion-binding motif and the altered motif. In Co(NH3)6(III) titrations, we observed large 1H and 31P chemical shift perturbations for the motif and found that outer-sphere complexation of Co(NH3)6(III) is possible for this motif. From the reinvestigation of our previous 15N chemical shift data for Cd(II) binding, in comparison with those of organometallic compounds, we conclude that Cd(II) can form an inner-sphere complex with the nucleobase in the motif. Therefore, the A9/G10.1 site was found to accept both inner-sphere and outer-sphere complexations. The Mg(II) titration for a slightly different motif from the A9/G10.1 site (G10.1-C11.1 to A10.1-U11.1) revealed that its affinity to Mg(II) was drastically reduced, although the ribozyme with this altered motif is known to retain enzymatic activities. This observation suggests that the metal ion at these motifs is not a catalytic center of hammerhead ribozymes.
Tanaka
Yoshiyuki
Y
Gene Function Research Center, National Institute of Advanced Industrial Science and Technology, Central 4, 1-1-1 Higashi, Tsukuba Science City 305-8562, Japan. tanaka@mail.pharm.tohoku.ac.jp
Kasai
Yasuhiro
Y
Mochizuki
Shunsuke
S
Wakisaka
Akihiro
A
Morita
Eugene H
EH
Kojima
Chojiro
C
Toyozawa
Atsushi
A
Kondo
Yoshinori
Y
Taki
Masumi
M
Takagi
Yasuomi
Y
Inoue
Atsushi
A
Yamasaki
Kazuhiko
K
Taira
Kazunari
K
D002412:Cations; D008670:Metals / Q000737:chemistry* / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000737:chemistry* / Q000378:metabolism*
D002412:Cations; D008670:Metals; D016337:RNA, Catalytic; C427204:hammerhead ribozyme