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PDBsum entry 2fyh
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References listed in PDB file
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Key reference
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Title
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Characterization of a heat-Stable enzyme possessing gtp-Dependent RNA ligase activity from a hyperthermophilic archaeon, Pyrococcus furiosus.
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Authors
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A.Kanai,
A.Sato,
Y.Fukuda,
K.Okada,
T.Matsuda,
T.Sakamoto,
Y.Muto,
S.Yokoyama,
G.Kawai,
M.Tomita.
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Ref.
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Rna, 2009,
15,
420-431.
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PubMed id
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Note: In the PDB file this reference is
annotated as "TO BE PUBLISHED". The citation details given above have
been manually determined.
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Abstract
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Using an expression protein library of a hyperthermophilic archaeon, Pyrococcus
furiosus, we identified a gene (PF0027) that encodes a protein with heat-stable
cyclic nucleotide phosphodiesterase (CPDase) activity. The PF0027 gene encoded a
21-kDa protein and an amino acid sequence that showed approximately 27% identity
to that of the 2'-5' tRNA ligase protein, ligT (20 kDa), from Escherichia coli.
We found that the purified PF0027 protein possessed GTP-dependent RNA ligase
activity and that synthetic tRNA halves bearing 2',3'-cyclic phosphate and 5'-OH
termini were substrates for the ligation reaction in vitro. GTP hydrolysis was
not required for the reaction, and GTPgammaS enhanced the tRNA ligation activity
of PF0027 protein, suggesting that the ligation step is regulated by a novel
mechanism. In comparison to the strong CPDase activity of the PF0027 protein,
the RNA ligase activity itself was quite weak, and the ligation product was
unstable during in vitro reaction. Finally, we used NMR to determine the
solution structure of the PF0027 protein and discuss the implications of our
results in understanding the role of the PF0027 protein.
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