Synthesis of calf prochymosin (prorennin) in Escherichia coli.
A gene for calf prochymosin (prorennin) has been reconstructed from chemically synthesized oligodeoxyribonucleotides and cloned DNA copies of preprochymosin mRNA. This gene has been inserted into a bacterial expression plasmid containing the Escherichia coli tryptophan promoter and a bacterial ribosome binding site. Induction of transcription from the tryptophan promoter results in prochymosin synthesis at a level of up to 5% of total protein. The enzyme has been purified from bacteria by extraction with urea and chromatography on DEAE-cellulose and converted to enzymatically active chymosin by acidification and neutralization. Bacterially produced chymosin is as effective in clotting milk as the natural enzyme isolated from calf stomach.
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D012085:Chymosin / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D004262:DNA Restriction Enzymes; D004792:Enzyme Precursors / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics
D004792:Enzyme Precursors; D012333:RNA, Messenger; D004247:DNA; D004262:DNA Restriction Enzymes; C032443:prorennin; D012085:Chymosin