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J. Biol. Chem., Vol. 276, Issue 43, 40041-40049, October 26, 2001
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,
§¶,
,
§, and
§
From the The CCA-adding enzyme (ATP:tRNA
adenylyltransferase or CTP:tRNA cytidylyltransferase (EC 2.7.7.25))
generates the conserved CCA sequence responsible for the attachment of
amino acid at the 3' terminus of tRNA molecules. It was shown that
enzymes from various organisms strictly recognize the elbow region of
tRNA formed by the conserved D- and T-loops. However, most of the
mammalian mitochondrial (mt) tRNAs lack consensus sequences in both D-
and T-loops. To characterize the mammalian mt CCA-adding enzymes, we
have partially purified the enzyme from bovine liver mitochondria and
determined cDNA sequences from human and mouse dbESTs by mass spectrometric analysis. The identified sequences contained typical amino-terminal peptides for mitochondrial protein import and had characteristics of the class II nucleotidyltransferase superfamily that
includes eukaryotic and eubacterial CCA-adding enzymes. The human
recombinant enzyme was overexpressed in Escherichia coli, and its CCA-adding activity was characterized using several mt tRNAs as
substrates. The results clearly show that the human mt CCA-adding enzyme can efficiently repair mt tRNAs that are poor substrates for the E. coli enzyme although both enzymes
work equally well on cytoplasmic tRNAs. This suggests that the
mammalian mt enzymes have evolved so as to recognize mt tRNAs with
unusual structures.
Department of Integrated Biosciences,
Graduate School of Frontier Sciences and § Department of
Chemistry and Biotechnology, Graduate School of Engineering, University
of Tokyo, Bldg. FSB-301, 5-1-5 Kashiwanoha, Kashiwa, Chiba Prefecture,
277-8562, Japan and
RIKEN Genomic Science Center, Suehiro-cho
1-7-22, Tsurumi-ku, Yokohama-shi, Kanagawa 230-0045, Japan
All cDNA sequences encoding mammalian mitochondrial CCA-adding enzymes reported in this paper have been submitted to the DDBJ/GenBankTM/EBI Data Bank with accession numbers AB063105, AB063106, and AB063107.
¶ To whom correspondence should be addressed. Tel.: 81-471-36-5401; Fax: 81-471-36-3602; E-mail: t-suzuki@k.u-tokyo.ac.jp.This article has been cited by other articles:
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