【作者】 肖睦； Michael J. Leibowitz； 张翼；

【Author】 ,Michael J. Leibowitz Department of Molecular Genetics, Microbiology and Immunology, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854-5635, and Cancer Institute of New Jersey, New Brunswick, New Jersey 08901, USA.

【机构】 武汉大学生命科学学院； Department of Molecular Genetics,Microbiology and Immunology,University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School,Piscataway,New Jersey 08854-5635 and Cancer Institute of New Jersey,New Brunswick,New Jersey 08901 USA.；

【摘要】 <正>Folding of the major population of Tetrahymena intron RNA to the catalytically active structure is trapped in a slow pathway. In this report, folding of Candida albicans intron was investigated using trans-acting Ca.L-11 ribozyme as a model. We demonstrated that both the catalytic activity (k_obs) and compact folding equilbrium of Ca.L-11 are strong¹., dependent on Mg²⁺ under the physiological concentrations, with both showing a Mg²⁺ Hill coefficient of 3. Formation of the compact structure of Ca.L-11 is shown to occur very rapidly, in a sub-second time scale similar to that of T1 cleavage. Most of the ribozyme RNA population folds to the catalytically active structure in a rate constant of 2 min^-1 at 10 mM Mg²⁺; neither slower kinetics or obvious Mg²⁺-inhibition is observed. These results suggest that folding of the Ca.L-11 ribozyme is initiated by a rapid magnesium-dependent RNA compaction, which is followed by a slower searching for the native contacts to form the catalytically active structure without the interference of the long-lived trapped states. This model thus provides an ideal system to address a range of interesting aspects of RNA folding, such as conformational searching, ion binding and the role of productive intermediates.更多还原

【Abstract】 Folding of the major population of Tetrahymena intron RNA to the catalytically active structure is trapped in a slow pathway. In this report, folding of Candida albicans intron was investigated using trans-acting Ca.L-11 ribozyme as a model. We demonstrated that both the catalytic activity (k_obs) and compact folding equilibrium of Ca.L-11 are strong., dependent on Mg²⁺ under the physiological concentrations, with both showing a Mg²⁺ Hill coefficient of 3. Formation of the compact structure of Ca.L- 11 is shown to occur very rapidly, in a sub-second time scale similar to that of T1 cleavage. Most of the ribozyme RNA population folds to the catalytically active structure in a rate constant of 2 min1 at 10 mM Mg²⁺; neither slower kinetics or obvious Mg²⁺-inhibition is observed. These results suggest that folding of the Ca.L-11 ribozyme is initiated by a rapid magnesium-dependent RNA compaction, which is followed by a slower searching for the native contacts to form the catalytically active structure without the interference of the long-lived trapped states. This model thus provides an ideal system to address a range of interesting aspects of RNA folding, such as conformational searching, ion binding and the role of productive intermediates.更多还原