Isolation and characterization of a NADP-dependent glutamate dehydrogenase gene from the primitive eucaryote Giardia lamblia.
Giardia lamblia is believed to be the earliest branching derivative from the eucaryotic lineage. Genomic and cDNA clones encoding the giardia NADP-dependent glutamate dehydrogenase have been isolated and characterized. Southern hydridization using genomic DNA indicates that the gene encoding this activity is unique and single copy. Primer extension, S1 nuclease protection, and genomic and cDNA sequence analysis demonstrate that gene transcripts are initiated within a conserved AT-rich sequence element immediately preceding the ATG translation initiation codon and the short 5' untranslated region is not extended by transsplicing. The open reading frame is 1350 nucleotides in length and encodes a protein of 449 amino acids. The reading frame is not interrupted by introns and the primary transcript is probably not subjected to RNA editing. In the strictly anaerobic metabolism of giardia, NADP-dependent glutamate dehydrogenase activity participates along with alanine aminotransferase, in the cyclic dissipation of reducing equivalents (NADPH) through the conversion of pyruvate to alanine. The deduced amino acid sequence of the giardia protein exhibits substantial homology to numerous fungal and eubacterial NADP-dependent glutamate dehydrogenases. Comparisons of alignment gap positions and amino acid identities indicate that the giardia sequence is at least as similar or more similar to the eubacterial sequence than it is to the fungal sequence. This supports the hypothesis that giardia diverged very early from the eucaryotic lineage.
Yee
J
J
Program in Evolutionary Biology, Canadian Institute for Advanced Research, Vancouver, British Columbia.
Dennis
P P
PP
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D016054:DNA, Protozoan / Q000235:genetics; D016829:Giardia lamblia / Q000201:enzymology*; D005969:Glutamate Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000235:genetics; D016053:RNA, Protozoan / Q000235:genetics; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D016054:DNA, Protozoan; D012333:RNA, Messenger; D016053:RNA, Protozoan; D005969:Glutamate Dehydrogenase