Separate metal requirements for loop interactions and catalysis in the extended hammerhead ribozyme.
An extended hammerhead ribozyme derived from Schistosoma mansoni, including conserved loops in stems I and II, has been examined to directly monitor the relationship between docking of loops and its activity using site-directed spin labeling (SDSL) and EPR spectroscopy. Dynamics with EPR spectroscopy and fast-quench kinetics measurements have shown that the docking of stems I and II occurs at low Mg2+ concentrations ([Mg2+]1/2,dock = 0.7 mM, 0.1 M NaCl), but a much weaker Mg2+ interaction ([Mg2+]1/2,cat approximately 90 mM) increases activity to very high maximum rates of approximately 1 s-1 at 0.1 M Na+ and pH 7.0.
Kim
Nak-Kyoon
NK
Department of Chemistry, Texas A&M University, College Station, TX 77843, USA.
Murali
Ayaluru
A
DeRose
Victoria J
VJ
D000818:Animals; D002384:Catalysis; D020134:Catalytic Domain; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry*; D016297:Mutagenesis, Site-Directed; D016337:RNA, Catalytic / Q000737:chemistry*; D043262:Ribonuclease P / Q000737:chemistry / Q000502:physiology; D012550:Schistosoma mansoni / Q000201:enzymology
D016337:RNA, Catalytic; C427204:hammerhead ribozyme; D043262:Ribonuclease P; D008274:Magnesium