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bioRxiv · 10.1101/666792

Kinetic mechanism of human mitochondrial RNase P

Liu, X.·Wu, N.·Shanmuganathan, A.·Klemm, B. P.·Howard, M. J.·Lim, W. H.·Koutmos, M.·Fierke, C. A.
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Abstract

A first step in processing mitochondrial precursor tRNA (pre-tRNA) is cleavage of the 5 leader catalyzed by ribonuclease P (RNase P). Human mitochondrial RNase P (mtRNase P) is composed of three protein subunits: mitochondrial RNase P protein (MRPP) 1, 2 and 3. Even though MRPP3 is the metallonuclease subunit responsible for catalysis, cleavage is observed only in the presence of the MRPP1/2 subcomplex. To understand the functional role of MRPP1/2, we reconstituted human mitochondrial RNase P in vitro and performed kinetic and thermodynamic analyses. MRPP1/2 significantly enhances both the catalytic activity and the apparent substrate affinity of mtRNase P. Additionally, pull-down and binding data demonstrate synergy between binding pre-tRNA and formation of a catalytically active MRPP1/2/3 complex. These data suggest that conformational changes in the MRPP1/2-pre-tRNA complex lead to protein-protein or protein-RNA interactions that increase both MRPP3 recognition and cleavage efficiency. This work presents the first kinetic model for human mtRNase P, providing a fundamental framework for the function of MRPP1/2 for recognition and processing of pre-tRNA.

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Liu, X., Wu, N., Shanmuganathan, A., Klemm, B. P., Howard, M. J., Lim, W. H., Koutmos, M., Fierke, C. A.. 2019-06-11. Kinetic mechanism of human mitochondrial RNase P. https://doi.org/10.1101/666792

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