Abstract
A human CAD cDNA encoding a trifunctional enzyme of carbamoylphosphate synthetase-aspartate transcarbamoylase-dihydroorotase, which catalyzes the first three steps of de novo pyrimidine nucleotide biosynthesis, was cloned from a human fibroblast cell line of TIG-1-20 by polymerase chain reaction (PCR). The predicted open reading frame encodes a protein of 2,225 amino acids with a deduced molecular weight (Mr) OF 242,913. The deduced amino acid sequence exhibits 95.3 and 76.1% identity with the CAD sequences of hamster and Squalus acanthias. The DNA fragment of 6,679 bp containing the full-length coding sequence was amplified by nested PCR using the first-strand cDNA of human cell lines of TIG-1-20 and COLO205 as a template. Southern blot analysis suggested that the CAD gene exists as a single copy in the human genome.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Animals
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Aspartate Carbamoyltransferase / biosynthesis*
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Aspartate Carbamoyltransferase / genetics
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Base Sequence
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Blotting, Southern
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Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing) / biosynthesis*
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Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing) / genetics
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Cell Line
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Cloning, Molecular
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Cricetinae
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DNA Primers
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DNA, Complementary
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Dihydroorotase / biosynthesis*
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Dihydroorotase / genetics
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Dogfish
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Fibroblasts
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Humans
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Molecular Sequence Data
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Molecular Weight
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Multienzyme Complexes / biosynthesis*
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Multienzyme Complexes / genetics
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Open Reading Frames
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Polymerase Chain Reaction
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Pyrimidines / biosynthesis
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Restriction Mapping
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Sequence Homology, Amino Acid
Substances
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CAD trifunctional enzyme
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DNA Primers
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DNA, Complementary
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Multienzyme Complexes
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Pyrimidines
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Aspartate Carbamoyltransferase
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Dihydroorotase
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Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing)