Fine structure analysis of the Chinese hamster AS gene encoding asparagine synthetase.
Overlapping cDNAs for Chinese hamster ovary (CHO) asparagine synthetase (AS) were isolated from a library prepared from an AS-overproducing cell line. The sequence was determined and shown to contain an open reading frame encoding a protein of Mr 64,300. The predicted amino acid sequence for the CHO AS enzyme was compared to that of the human AS enzyme and found to be 95% homologous. A potential glutamine amide transfer domain, with sequence similarity to amidotransferases from bacteria and yeast, was identified in the N-terminal portion of the protein. The cDNAs were used to screen a library of phage containing wild type CHO DNA and the genomic AS sequences were detected on three overlapping phages. Determination of the fine structural organization showed that the CHO AS gene spanned 19 kilobases and was composed of 12 exons, three of which contained the glutamine amidotransferase domain. The 5' flanking sequences were highly G + C-rich and, like other housekeeping genes, lacked TATA and CAAT boxes.
Andrulis
I L
IL
Department of Research, Mt. Sinai Hospital, Toronto, Ontario, Canada.
Shotwell
M
M
Evans-Blackler
S
S
Zalkin
H
H
Siminovitch
L
L
Ray
P N
PN
D000595:Amino Acid Sequence; D000818:Animals; D001217:Aspartate-Ammonia Ligase / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D015139:Blotting, Southern; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D004274:DNA, Recombinant; D005786:Gene Expression Regulation; D005973:Glutamine / Q000378:metabolism; D006801:Humans; D008025:Ligases / Q000235:genetics*; D008969:Molecular Sequence Data; D012313:RNA / Q000302:isolation & purification; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014166:Transferases / Q000235:genetics / Q000378:metabolism
D004274:DNA, Recombinant; D005973:Glutamine; D012313:RNA; D004247:DNA; D014166:Transferases; D008025:Ligases; D001217:Aspartate-Ammonia Ligase