Cleavage Mediated by the Catalytic Domain of Bacterial RNase P RNA |
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| Authors: | Wu Shiying Kikovska Ema Lindell Magnus Kirsebom Leif A |
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| Institution: | 3. Committee on Microbiology, University of Chicago, Chicago, Illinois, 60637;5. Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois 60637;4. Howard T. Ricketts Laboratory, Argonne National Laboratory, Argonne, Illinois 60439;6. Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721;1. State Key Laboratory of Soil Erosion and Dryland Farming on the Loess Plateau, Institute of Soil and Water Conservation, Northwest A&F University, Yangling, 712100, China;2. University of Chinese Academy of Sciences, Beijing, 100049, China;3. College of Xingzhi, Zhejiang Normal University, Jinhua, 321000, China;4. CAS Center for Excellence in Quaternary Science and Global Change, Chinese Academy of Sciences, Xian, 710061, China;1. NNF-CFB, Technical University of Denmark, 2800 Kongens Lyngby, Denmark;1. Department of Microbial Pathogenesis, Yale University School of Medicine, New Haven, CT 06536, USA;2. Microbial Sciences Institute, Yale University, West Haven, CT 06516, USA;3. Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06511, USA |
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| Abstract: | Like other RNA molecules, RNase P RNA (RPR) is composed of domains, and these have different functions. Here, we provide data demonstrating that the catalytic (C) domain of Escherichia coli (Eco) RPR when separated from the specificity (S) domain mediates cleavage using various model RNA hairpin loop substrates. Compared to full-length Eco RPR, the rate constant, k(obs), of cleavage for the truncated RPR (CP RPR) was reduced 30- to 13,000-fold depending on substrate. Specifically, the structural architecture of the -1/+73 played a significant role where a C(-1)/G(+73) pair had the most dramatic effect on k(obs). Substitution of A(248) (E. coli numbering), positioned near the cleavage site in the RNase P-substrate complex, with G in the CP RPR resulted in 30-fold improvement in rate. In contrast, strengthening the interaction between the RPR and the 3' end of the substrate only had a modest effect. Interestingly, although deleting the S-domain gave a reduction in the rate, it resulted in a less erroneous RPR with respect to cleavage site selection. These data support and extend our understanding of the coupling between the distal interaction between the S-domain and events at the active site. Our findings will also be discussed with respect to the structure of RPR derived from different organisms. |
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