Publications

• Closely related CC- and A-adding enzymes collaborate to construct and repair the 3'-terminal CCA of tRNA in Synechocystis sp. and Deinococcus radiodurans.

  Tomita K., Weiner A.M.

  The 3'-terminal CCA sequence of tRNA is faithfully constructed and repaired by the CCA-adding enzyme (ATP(CTP):tRNA nucleotidyltransferase) using CTP and ATP as substrates but no nucleic acid template. Until recently, all CCA-adding enzymes from all three kingdoms appeared to be composed of a sing ... >> More

  The 3'-terminal CCA sequence of tRNA is faithfully constructed and repaired by the CCA-adding enzyme (ATP(CTP):tRNA nucleotidyltransferase) using CTP and ATP as substrates but no nucleic acid template. Until recently, all CCA-adding enzymes from all three kingdoms appeared to be composed of a single kind of polypeptide with dual specificity for adding both CTP and ATP; however, we recently found that in Aquifex aeolicus, which lies near the deepest root of the eubacterial 16 S rRNA-based phylogenetic tree, CCA addition represents a collaboration between closely related CC-adding and A-adding enzymes (Tomita, K. and Weiner, A. M. (2001) Science 294, 1334-1336). Here we show that in Synechocystis sp. and Deinococcus radiodurans, as in A. aeolicus, CCA is added by homologous CC- and A-adding enzymes. We also find that the eubacterial CCA-, CC-, and A-adding enzymes, as well as the related eubacterial poly(A) polymerases, each fall into phylogenetically distinct groups derived from a common ancestor. Intriguingly, the Thermatoga maritima CCA-adding enzyme groups with the A-adding enzymes, suggesting that these distinct tRNA nucleotidyltransferase activities can intraconvert over evolutionary time. << Less

  J. Biol. Chem. 277:48192-48198(2002) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Collaboration between CC- and A-adding enzymes to build and repair the 3'-terminal CCA of tRNA in Aquifex aeolicus.

  Tomita K., Weiner A.M.

  The universal 3'-terminal CCA sequence of all transfer RNAs (tRNAs) is repaired, and sometimes constructed de novo, by the CCA-adding enzyme [ATP(CTP):tRNA nucleotidyltransferase]. This RNA polymerase has no nucleic acid template, yet faithfully builds the CCA sequence one nucleotide at a time usi ... >> More

  The universal 3'-terminal CCA sequence of all transfer RNAs (tRNAs) is repaired, and sometimes constructed de novo, by the CCA-adding enzyme [ATP(CTP):tRNA nucleotidyltransferase]. This RNA polymerase has no nucleic acid template, yet faithfully builds the CCA sequence one nucleotide at a time using cytidine triphosphate (CTP) and adenosine triphosphate (ATP) as substrates. All previously characterized CCA-adding enzymes from all three kingdoms are single polypeptides with CCA-adding activity. Here, we demonstrate through biochemical and genetic approaches that CCA addition in Aquifex aeolicus requires collaboration between two related polypeptides, one that adds CC and another that adds A. << Less

  Science 294:1334-1336(2001) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Geobacter sulfurreducens contains separate C- and A-adding tRNA nucleotidyltransferases and a poly(A) polymerase.

  Bralley P., Cozad M., Jones G.H.

  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 ... >> More

  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. << Less

  J. Bacteriol. 191:109-114(2009) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Evolution of tRNA nucleotidyltransferases: a small deletion generated CC-adding enzymes.

  Neuenfeldt A., Just A., Betat H., Moerl M.

  CCA-adding enzymes are specialized polymerases that add a specific sequence (C-C-A) to tRNA 3' ends without requiring a nucleic acid template. In some organisms, CCA synthesis is accomplished by the collaboration of evolutionary closely related enzymes with partial activities (CC and A addition). ... >> More

  CCA-adding enzymes are specialized polymerases that add a specific sequence (C-C-A) to tRNA 3' ends without requiring a nucleic acid template. In some organisms, CCA synthesis is accomplished by the collaboration of evolutionary closely related enzymes with partial activities (CC and A addition). These enzymes carry all known motifs of the catalytic core found in CCA-adding enzymes. Therefore, it is a mystery why these polymerases are restricted in their activity and do not synthesize a complete CCA terminus. Here, a region located outside of the conserved motifs was identified that is missing in CC-adding enzymes. When recombinantly introduced from a CCA-adding enzyme, the region restores full CCA-adding activity in the resulting chimera. Correspondingly, deleting the region in a CCA-adding enzyme abolishes the A-incorporating activity, also leading to CC addition. The presence of the deletion was used to predict the CC-adding activity of putative bacterial tRNA nucleotidyltransferases. Indeed, two such enzymes were experimentally identified as CC-adding enzymes, indicating that the existence of the deletion is a hallmark for this activity. Furthermore, phylogenetic analysis of identified and putative CC-adding enzymes indicates that this type of tRNA nucleotidyltransferases emerged several times during evolution. Obviously, these enzymes descend from CCA-adding enzymes, where the occurrence of the deletion led to the restricted activity of CC addition. A-adding enzymes, however, seem to represent a monophyletic group that might also be ancestral to CCA-adding enzymes. Yet, experimental data indicate that it is possible that A-adding activities also evolved from CCA-adding enzymes by the occurrence of individual point mutations. << Less

  Proc. Natl. Acad. Sci. U.S.A. 105:7953-7958(2008) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Schizosaccharomyces pombe contains separate CC- and A-adding tRNA nucleotidyltransferases.

  Reid N.E., Ngou J.S., Joyce P.B.M.

  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 ... >> More

  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. The fungus, Schizosaccharomyces pombe, has two genes (cca1 and cca2) that are thought, based on predicted amino acid sequences, to encode tRNA nucleotidyltransferases. Here, we show that both genes together are required to complement a Saccharomyces cerevisiae strain bearing a null mutation in the single gene encoding its tRNA nucleotidyltransferase. Using enzyme assays we show further that the purified S. pombe cca1 gene product specifically adds two cytidine residues to a tRNA substrate lacking this sequence while the cca2 gene product specifically adds the terminal adenosine residue thereby completing the CCA sequence. These data indicate that S. pombe represents the first eukaryote known to have separate CC- and A-adding activities for tRNA maturation and repair. In addition, we propose that a novel structural change in a tRNA nucleotidyltransferase is responsible for defining a CC-adding enzyme. << Less

  Biochem. Biophys. Res. Commun. 508:785-790(2019) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.