The genome of Geobacter sulfurreducens contains three genes whose sequences are quite similar to sequences encoding known members of an RNA nucleotidyltransferase superfamily that includes tRNA nucleotidyltransferases and poly(A) polymerases. Reverse transcription-PCR using G. sulfurreducens total RNA demonstrated that the genes encoding these three proteins are transcribed. These genes, encoding proteins designated NTSFI, NTSFII, and NTSFIII, were cloned and overexpressed in Escherichia coli. The corresponding enzymes were purified and assayed biochemically, resulting in identification of NTSFI as a poly(A) polymerase, NTSFII as a C-adding tRNA nucleotidyltransferase, and NTSFIII as an A-adding tRNA nucleotidyltransferase. Analysis of G. sulfurreducens rRNAs and mRNAs revealed the presence of heteropolymeric RNA 3' tails. This is the first characterization of a bacterial system that expresses separate C- and A-adding tRNA nucleotidyltransferases and a poly(A) polymerase.
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http://dx.doi.org/10.1128/JB.01166-08 | DOI Listing |
Front Microbiol
April 2024
Department of Science, Università degli Studi della Basilicata, Potenza, Italy.
Mol Biol Evol
March 2021
Institute for Biochemistry, Leipzig University, Leipzig, Germany.
tRNAs are important players in the protein synthesis machinery, where they act as adapter molecules for translating the mRNA codons into the corresponding amino acid sequence. In a series of highly conserved maturation steps, the primary transcripts are converted into mature tRNAs. In the amoebozoan Acanthamoeba castellanii, a highly unusual evolution of some of these processing steps was identified that are based on unconventional RNA polymerase activities.
View Article and Find Full Text PDFInt J Mol Sci
January 2020
Institute for Biochemistry, Leipzig University, Brüderstraße 34, 04103 Leipzig, Germany.
Synthesis of the CCA end of essential tRNAs is performed either by CCA-adding enzymes or as a collaboration between enzymes restricted to CC- and A-incorporation. While the occurrence of such tRNA nucleotidyltransferases with partial activities seemed to be restricted to Bacteria, the first example of such split CCA-adding activities was reported in . Here, we demonstrate that the choanoflagellate also carries CC- and A-adding enzymes.
View Article and Find Full Text PDFNucleic Acids Res
April 2019
Institute for Biochemistry, Leipzig University, Brüderstr. 34, 04103 Leipzig, Germany.
Correct synthesis and maintenance of functional tRNA 3'-CCA-ends is a crucial prerequisite for aminoacylation and must be achieved by the phylogenetically diverse group of tRNA nucleotidyltransferases. While numerous reports on the in vitro characterization exist, robust analysis under in vivo conditions is lacking. Here, we utilize Escherichia coli RNase T, a tRNA-processing enzyme responsible for the tRNA-CCA-end turnover, to generate an in vivo system for the evaluation of A-adding activity.
View Article and Find Full Text PDFBiochem Biophys Res Commun
January 2019
Department of Chemistry and Biochemistry, Centre for Structural and Functional Genomics, Concordia University, 7141 Sherbrooke St. W, Montréal, Québec, H4B 1R6, Canada. Electronic address:
A specific cytidine-cytidine-adenosine (CCA) sequence is required at the 3'-terminus of all functional tRNAs. This sequence is added during tRNA maturation or repair by tRNA nucleotidyltransferase enzymes. While most eukaryotes have a single enzyme responsible for CCA addition, some bacteria have separate CC- and A-adding activities.
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