Internal equilibrium of the hammerhead ribozyme is altered by the length of certain covalent cross-links.
A method was developed that permits covalent cross-links of different linker lengths to be introduced into RNA at defined positions. The previous observation that a cross-link between stems I and II of the hammerhead ribozyme was confirmed and further explored. By examining the catalytic consequences of varying the position and length of this cross-link, we conclude that the previously proposed conformational dampening model cannot sufficiently explain the increase in ligation rate induced by the cross-link. Rather, the cross-link constrains the cleaved hammerhead into a structure that more closely resembles the transition state, thereby increasing the reverse ligation rate relative to a non-cross-linked control.
Blount
Kenneth F
KF
Department of Chemistry and Biochemistry, University of Colorado-Boulder, UCB 215, Boulder, CO 80309, USA.
Uhlenbeck
Olke C
OC
D003432:Cross-Linking Reagents / Q000737:chemistry*; D008025:Ligases / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000737:chemistry / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism*; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D016337:RNA, Catalytic; C427204:hammerhead ribozyme; D012260:Ribonucleases; D008025:Ligases