Mechanism for the definition of elongation and termination by the class II CCA-adding enzyme

Yukimatsu Toh, Daijiro Takeshita, Tomoyuki Numata, Shuya Fukai, Osamu Nureki, Kozo Tomita

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

The CCA-adding enzyme synthesizes the CCA sequence at the 3′ end of tRNA without a nucleic acid template. The crystal structures of class II Thermotoga maritima CCA-adding enzyme and its complexes with CTP or ATP were determined. The structure-based replacement of both the catalytic heads and nucleobase-interacting neck domains of the phylogenetically closely related Aquifex aeolicus A-adding enzyme by the corresponding domains of the T. maritima CCA-adding enzyme allowed the A-adding enzyme to add CCA in vivo and in vitro. However, the replacement of only the catalytic head domain did not allow the A-adding enzyme to add CCA, and the enzyme exhibited (A, C)-adding activity. We identified the region in the neck domain that prevents (A, C)-adding activity and defines the number of nucleotide incorporations and the specificity for correct CCA addition. We also identified the region in the head domain that defines the terminal A addition after CC addition. The results collectively suggest that, in the class II CCA-adding enzyme, the head and neck domains collaboratively and dynamically define the number of nucleotide additions and the specificity of nucleotide selection.

Original languageEnglish
Pages (from-to)3353-3365
Number of pages13
JournalEMBO Journal
Volume28
Issue number21
DOIs
Publication statusPublished - Nov 1 2009

Fingerprint

Elongation
Head
Thermotoga maritima
Neck
Nucleotides
Cytidine Triphosphate
Transfer RNA
Nucleic Acids
Catalytic Domain
Adenosine Triphosphate
Crystal structure
tRNA nucleotidyltransferase
Enzymes
A adding enzyme

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

Mechanism for the definition of elongation and termination by the class II CCA-adding enzyme. / Toh, Yukimatsu; Takeshita, Daijiro; Numata, Tomoyuki; Fukai, Shuya; Nureki, Osamu; Tomita, Kozo.

In: EMBO Journal, Vol. 28, No. 21, 01.11.2009, p. 3353-3365.

Research output: Contribution to journalArticle

Toh, Yukimatsu ; Takeshita, Daijiro ; Numata, Tomoyuki ; Fukai, Shuya ; Nureki, Osamu ; Tomita, Kozo. / Mechanism for the definition of elongation and termination by the class II CCA-adding enzyme. In: EMBO Journal. 2009 ; Vol. 28, No. 21. pp. 3353-3365.
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