Identification of the alpha-aminoadipic semialdehyde dehydrogenase-phosphopantetheinyl transferase gene, the human ortholog of the yeast LYS5 gene.
In mammals, L-lysine is first catabolized to alpha-aminoadipate semialdehyde by the bifunctional enzyme alpha-aminoadipate semialdehyde synthase (AASS), followed by a conversion to alpha-aminoadipate by alpha-aminoadipate semialdehyde dehydrogenase. In Saccharomyces cerevisiae, which synthesize rather than degrade lysine, the latter activity requires two distinct genes. LYS2 encodes the alpha-aminoadipate reductase activity, while LYS5 encodes a phosphopantetheinyl transferase activity that is required to activate Lys2p. We have identified a full-length human cDNA homologous to the yeast LYS5 gene. The cDNA contains an open-reading frame of 930 bp predicted to encode 309 amino acids, and the human protein is 26% identical and 44% similar to its yeast counterpart. In Northern blot analysis the cDNA hybridizes to a single transcript of approximately 3 kb in all tissues except testis, where there is an additional transcript of 1.5 kb. Expression is highest in brain followed by heart and skeletal muscle, and to a lesser extent in liver. We further identified three human genomic BAC clones containing the human gene. Fluorescence in situ hybridization (FISH) analysis using the BAC clones mapped the gene to chromosome 11q22 while alignment of the cDNA and genomic sequences allowed partial identification of the intron-exon boundaries. Finally, using one-step homologous recombination in S. cerevisiae we generated a lys5 knockout strain. Complementation studies in the yeast knockout demonstrate that the human homolog encodes alpha-aminoadipate dehydrogenase phosphopantetheinyl transferase activity. We hypothesize that defects in this gene may result in pipecolic acidemia.
Praphanphoj
V
V
Department of Pediatrics, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Sacksteder
K A
KA
Gould
S J
SJ
Thomas
G H
GH
Geraghty
M T
MT
D000445:Aldehyde Oxidoreductases / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002874:Chromosome Mapping; D002880:Chromosomes, Human, Pair 11 / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000032:analysis / Q000302:isolation & purification; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics*; D015870:Gene Expression; D005800:Genes, Fungal / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D050647:L-Aminoadipate-Semialdehyde Dehydrogenase; D008239:Lysine / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D017855:Transferases (Other Substituted Phosphate Groups) / Q000235:genetics*
D017931:DNA Primers; D018076:DNA, Complementary; D005656:Fungal Proteins; D000445:Aldehyde Oxidoreductases; D050647:L-Aminoadipate-Semialdehyde Dehydrogenase; D017855:Transferases (Other Substituted Phosphate Groups); C021364:holo-(acyl-carrier-protein) synthase; D008239:Lysine