Proceedings of the National Academy of Sciences · 1982 · 23 citations · 23 references
Life SciencesBiotechnologyMolecular BiologyGenetic EngineeringHormone BiochemistryProtein EngineeringHormone SecretionPancreatic HormonesPlasmid BiologyPlasmid EngineeringPlasmid GeneticsProtein Expression EngineeringEscherichia ColiGastrointestinal Hormone SecretionIntestinal Hormone SecretionPancreatic Beta-cell SecretionPancreatic Hormone SecretionPancreatic Islet SecretionPancreatic SecretionPlasmid Integration
A synthetic gene for porcine secretin was ligated with Pst I-cleaved pBR322 by its flanking synthetic Pst I linkers to produce a fused protein consisting of an amino-terminal portion of beta-lactamase and an entire secretin molecule. The hybrid plasmid was transferred into competent Escherichia coli cells. The plasmids, which were proved in our investigation to contain the secretin gene in the desired orientation, were then screened for the production of secretin. This was shown by radioimmunoassay and gel electrophoretic analysis of the polypeptides that were synthesized in E. coli minicells transformed with the hybrid plasmids. Secretin so produced, whose carboxy terminal residue may not be amidated in contrast with natural porcine secretin, showed the same activity in stimulating pancreatic secretion in a bioassay with anesthetized rats as does natural secretin.
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