Cloning, sequencing, and expression of a cDNA encoding beta-alanine synthase from rat liver.
A cDNA for beta-alanine synthase from rat liver has been isolated, sequenced, and characterized. beta-Alanine synthase clones were isolated from rat liver cDNA libraries in lambda gt11, using affinity-purified polyclonal antibodies against beta-alanine synthase protein. beta-Alanine synthase protein was not expressed with equal efficiency by all clones. One of the expressed fusion proteins has normal specific enzyme activity, and a second has reduced specific activity. Both clones were completely sequenced and yielded identical DNA sequence, except that one clone contained an additional 36 bases of 5' sequence. The various clones of this cDNA code for an EcoRI insert of 1.5 +/- 0.1 kb, and the open reading frame corresponds to a protein of 393 amino acids (M(r) = 44,042), in good agreement with the M(r) of approximately 42,000 for the native enzyme on SDS-gel electrophoresis. An 11-amino acid sequence was obtained from a tryptic peptide of native beta-alanine synthase; 11 codons for these same amino acids were found at the expected site in the sequenced cDNA, and confirm the open reading frame of the beta-alanine synthase cDNA. Chemical analysis of the native enzyme shows 2 zinc atoms per subunit, and the sequence of beta-alanine synthase contains 2 putative zinc-binding site motifs. Comparison of amino acid sequence, deduced from the cDNA sequence, to sequences in the protein data base showed that it is a unique sequence and that it has about 20% identity to aspartate carbamoyltransferase, ornithine carbamoyltransferase, urease, and leucine aminopeptidase; enzymes that bind comparable ligands or have a similar mechanism.
Kvalnes-Krick
K L
KL
Department of Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill 27599-7260.
Traut
T W
TW
D000581:Amidohydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA; D004926:Escherichia coli; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D051381:Rats; D015032:Zinc / Q000378:metabolism
D004247:DNA; D000581:Amidohydrolases; C021590:beta-ureidopropionase; D015032:Zinc