Expression of chemically synthesized alpha-neo-endorphin gene fused to E. coli alkaline phosphatase.
An alpha-neo-endorphin (alpha NE) gene, which we previously synthesized chemically and inserted into E. coli beta-galactosidase gene of pK013 plasmid, has been excised and fused to E. coli alkaline phosphatase (APase) gene. One of the transformants was named E15/pA alpha NE1. Under the APase gene regulation, APase-alpha NE chimeric protein was expressed at 1.3 X 10(6) molecules per cell, and accounted for about 60% of total cellular proteins. The HPLC pattern of CNBr treated E15/pA alpha NE1 was very simple reflecting the high content of the chimeric protein and low numbers of methionine residues in it. A series of genes encoding APase-alpha NE chimeric proteins in which 30 to 94 C-terminal amino acid residues were replaced by (met)-alpha NE, was cloned in E. coli. Transportation of the chimeric proteins to periplasmic space was studied. All chimeric proteins were apparently processed by signal peptidase but few, if any, was transported to the periplasmic space.
D000469:Alkaline Phosphatase / Q000235:genetics*; D000595:Amino Acid Sequence; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant / Q000378:metabolism; D004723:Endorphins / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005796:Genes; D005798:Genes, Bacterial; D007700:Kinetics; D010957:Plasmids; D011498:Protein Precursors / Q000235:genetics* / Q000302:isolation & purification; D001616:beta-Galactosidase / Q000235:genetics
D004274:DNA, Recombinant; D004723:Endorphins; D011498:Protein Precursors; C025681:alpha-neoendorphin; D004262:DNA Restriction Enzymes; D000469:Alkaline Phosphatase; D001616:beta-Galactosidase