Prolyl-tRNA synthetase from Thermus thermophilus is eukaryotic-like but aminoacylates prokaryotic tRNAPro
Aim. Cloning, sequencing and expression of the gene encoding prolyl-tRNA synthetase, a class IIa enzyme, from the extreme thermophile T. thermophilus HB8 (ProRSTT). Methods. Search for the ProRSTT gene was performed by Southern blot hybridization with chromosomal DNA, the digoxigenin-labeled PCR fr...
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| Date: | 2012 |
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| Main Authors: | , , |
| Format: | Article |
| Language: | English |
| Published: |
Інститут молекулярної біології і генетики НАН України
2012
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| Series: | Вiopolymers and Cell |
| Subjects: | |
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| Journal Title: | Digital Library of Periodicals of National Academy of Sciences of Ukraine |
| Cite this: | Prolyl-tRNA synthetase from Thermus thermophilus is eukaryotic-like but aminoacylates prokaryotic tRNAPro / A.D. Yaremchuk, K.S. Boyarshin, M.A. Tukalo // Вiopolymers and Cell. — 2012. — Т. 28, № 6. — С. 434-440. — Бібліогр.: 22 назв. — англ. |
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Digital Library of Periodicals of National Academy of Sciences of Ukraine| Summary: | Aim. Cloning, sequencing and expression of the gene encoding prolyl-tRNA synthetase, a class IIa enzyme, from
the extreme thermophile T. thermophilus HB8 (ProRSTT). Methods. Search for the ProRSTT gene was performed by Southern blot hybridization with chromosomal DNA, the digoxigenin-labeled PCR fragments of DNA
being used as a probe. Results. The gene of T. thermophilus HB8 ProRS has been cloned and sequenced. The predicted 477-amino acid protein is significantly more similar in sequence to eukaryotic and archaeal than to eubacterial ProRS. Sequence analysis shows two distinct structural groups of ProRS which most likely had diverged
early in evolution: (1) eukaryotic/archaeal group characterized by the absence of an insertion domain between
motifs 2 and 3 and by the presence of an extra C-terminal domain beyond the normal class IIa anticodon binding
domain; and (2) prokaryotic with a very large insertion between motif 2 and 3 and no extra C-terminal domain.
Conclusions. T. thermophilus proS gene was overexpressed in Escherichia coli and overproduced ProRSTT was
purified to high homogeneity. In spite of its eukaryotic-like features, T. thermophilus ProRS exhibited highly
specific cross-species aminoacylation. The charging ability of the ProRSTT is restricted to prokaryotic tRNAPro.
Keywords: prolyl-tRNA synthetase from Thermus thermophilus, proS gene, сloning, sequencing, expression. |
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