Nuclear Protein-Only Ribonuclease P2 Structure and Biochemical Characterization Provide Insight into the Conserved Properties of tRNA 5' End Processing Enzymes

Karasik, Agnes; Shanmuganathan, Aranganathan; Howard, Michael J.; Fierke, Carol A.; Koutmos, Markos

doi:10.1016/j.jmb.2015.11.025

Nuclear Protein-Only Ribonuclease P2 Structure and Biochemical Characterization Provide Insight into the Conserved Properties of tRNA 5' End Processing Enzymes

Journal Article · Wed Dec 02 23:00:00 EST 2015 · Journal of Molecular Biology

DOI:https://doi.org/10.1016/j.jmb.2015.11.025· OSTI ID:1237761

Karasik, Agnes ^[1]; Shanmuganathan, Aranganathan ^[1]; Howard, Michael J. ^[2]; Fierke, Carol A. ^[2]; Koutmos, Markos ^[1]

Uniformed Services Univ. of the Health Sciences, Bethesda, MD (United States)
Univ. of Michigan, Ann Arbor, MI (United States)

Protein-only RNase Ps (PRORPs) are a recently discovered class of RNA processing enzymes that catalyze maturation of the 5' end of precursor tRNAs in Eukaryotes. PRORPs are found in the nucleus and/or organelles of most eukaryotic organisms. Arabidopsis thaliana is a representative organism that contains PRORP enzymes (PRORP1, PRORP2 and PRORP3) in both its nucleus and its organelles; PRORP2 and PRORP3 localize to the nucleus and PRORP1 localizes to the chloroplast and the mitochondria. Apart from their identification, almost nothing is known about the structure and function of PRORPs that act in the nucleus. Here, we use a combination of biochemical assays and X-ray crystallography to characterize A. thaliana PRORP2. We solved the crystal structure of PRORP2 (3.2 Å) revealing an overall V-shaped protein and conserved metallonuclease active-site structure. Our biochemical studies indicate that PRORP2 requires Mg²⁺ for catalysis and catalyzes the maturation of nuclear encoded substrates up to 10-fold faster than mitochondrial encoded precursor nad6 t-element under single-turnover conditions. We also demonstrate that PRORP2 preferentially binds precursor tRNAs containing short 5' leaders and 3' trailers; however, leader and trailer lengths do not significantly alter the observed rate constants of PRORP2 in single-turnover cleavage assays. In conclusion, our data provide a biochemical and structural framework to begin understanding how nuclear localized PRORPs recognize and cleave their substrates.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States)

Sponsoring Organization:: National Inst. of Health

OSTI ID:: 1237761

Journal Information:: Journal of Molecular Biology, Journal Name: Journal of Molecular Biology Journal Issue: 1 Vol. 428; ISSN 0022-2836

Publisher:: ElsevierCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

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Nuclear Protein-Only Ribonuclease P2 Structure and Biochemical Characterization Provide Insight into the Conserved Properties of tRNA 5' End Processing Enzymes

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