Characterization of cDNA encoding the human tRNA-guanine transglycosylase (TGT) catalytic subunit.
Queuosine (Q) is a 7-deazaguanosine found in the first position of the anticodon of tRNAs that recognize NAU and NAC codons (Tyr, Asn, Asp and His). Eukaryotes synthesize Q by the base-for-base exchange of queuine (Q base) for guanine in the unmodified tRNA, a reaction catalyzed by TGT. A search of the human EST database for sequences with significant homology to the well studied TGT from Escherichia coli identified several candidates for full-length (1.3-1.4 kb) cDNA clones. Three candidate cDNA clones, available from IMAGE Consortium, LLNL, (Lennon et al., 1996, Genomics 33, 151-152) were obtained: IMAGE Clone Id Nos. 611146, 1422928, and 72154. Here we report the complete sequences of these clones. IMAGE:72154 contains an ORF encoding a 44 kDa polypeptide with high homology to bacterial TGTs and was subcloned into the mammalian expression vector pMAMneo-Cat. When this construct was transfected into the TGT-negative cell line, GC(3)/c1 (Gündüz et al., 1992, Biochim. Biophys. Acta 1139, 229-238), it restored the ability of the cells to form Q-containing tRNA. This TGT cDNA sequence is encoded in human chromosome 19 clone CTC-539A10 (GenBank accession no. AC011475), enabling determination of the exon-intron boundaries for the TGT gene. The sequence of IMAGE:611146 is 5'-truncated by 76 bp compared to that from IMAGE:72154 and, except for two differences in the 3'-non-coding region, the remainder of the sequence is identical to that of IMAGE:72154. IMAGE:1422928 is a 1390 bp chimera: the 5'-portion, bp 1-708, is identical to a genomic DNA sequence from chromosome 15 (GenBank accession no. AC067805, bp 148976-149683); the 3'-end, bp 726-1390, is identical to the 3'-end of the TGT cDNA sequence from IMAGE:611146.
Deshpande
K L
KL
Department of Molecular Sciences, The University of Tennessee Health Science Center, 858 Madison Avenue, 38163, Memphis, TN, USA.
Katze
J R
JR
D001483:Base Sequence; D020134:Catalytic Domain; D002874:Chromosome Mapping; D002888:Chromosomes, Human, Pair 19 / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D005091:Exons; D005796:Genes / Q000235:genetics; D006147:Guanine / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D010430:Pentosyltransferases / Q000235:genetics* / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D006147:Guanine; C019549:queuine; D010430:Pentosyltransferases; C020837:queuine tRNA-ribosyltransferase