Coexpression of the cys E and cys M genes of Salmonella typhimurium in mammalian cells: a step towards establishing cysteine biosynthesis in sheep by transgenesis.
The Salmonella typhimurium genes for serine acetyltransferase (cys E) and O-acetylserine sulphydrylase B (cys M) were isolated and characterized in order to express these as transgenes in sheep to establish a cysteine biosynthesis pathway and, thereby, to achieve an increased rate of wool growth. Comparison of the S. typhimurium and Escherichia coli genes showed considerable homology, both at the nucleotide and amino acid sequence levels. The in vitro and in vivo expression studies showed that both genes could be transcribed and translated in eukaryotic cells and that their products could function as active enzymes. The cys M gene of S. typhimurium possessed a GUG initiation codon, like its E. coli counterpart, but translation could be initiated using this codon in eukaryotic cells to give an active enzyme product. Chinese hamster ovary cells, stably transfected with a tandem arrangement of the two genes, showed a capacity to synthesize cysteine in vivo, indicating the establishment of a cysteine biosynthesis pathway in these cells. The measured levels of activity of the gene products suggest that improved wool growth is possible by transgenesis of sheep with these genes.
Sivaprasad
A V
AV
Biochemistry Department, University of Adelaide, Australia.
Kuczek
E S
ES
Bawden
C S
CS
Rogers
G E
GE
D000123:Acetyltransferases / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D003545:Cysteine / Q000096:biosynthesis*; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D015698:Genomic Library; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D014176:Protein Biosynthesis; D011817:Rabbits; D015183:Restriction Mapping; D012156:Reticulocytes / Q000378:metabolism; D012486:Salmonella typhimurium / Q000201:enzymology* / Q000235:genetics*; D051050:Serine O-Acetyltransferase; D012756:Sheep / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection
D001426:Bacterial Proteins; C062886:CysB protein, Bacteria; C489265:CysB protein, Salmonella typhimurium; D004269:DNA, Bacterial; D009838:Oligodeoxyribonucleotides; D000123:Acetyltransferases; D051050:Serine O-Acetyltransferase; D003545:Cysteine