Determinants for protein localization: beta-lactamase signal sequence directs globin across microsomal membranes.
A hybrid gene containing 182 codons of Escherichia coli beta-lactamase at the amino terminus of the corresponding protein and 141 codons of alpha-globin at the carboxyl terminus was generated by inserting chimpanzee alpha-globin cDNA into the Pst I site of plasmid pBR322. RNA transcribed in vitro from this plasmid gave a corresponding hybrid protein in a wheat germ cell-free translation system. The hybrid protein was protected from tryptic digestion and the pre-beta-lactamase signal peptide was removed when dog pancreas membrane vesicles were present during translation. A deletion mutant containing 23 codons of pre-beta-lactamase signal sequence and 5 codons of mature beta-lactamase fused to the alpha-globin cDNA gave a shorter hybrid protein that behaved similarly. However, a mutation that removed essentially all of the pre-beta-lactamase sequence gave a protein that was neither protected nor processed. Hence, at most, only the signal peptide and the first 5 amino acids of beta-lactamase were necessary to convert alpha-globin (a cytoplasmic protein) into a secretory protein.
D000818:Animals; D002451:Cell Compartmentation; D004274:DNA, Recombinant; D005914:Globins / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism*; D002679:Pan troglodytes; D010455:Peptides / Q000502:physiology*; D011498:Protein Precursors / Q000502:physiology*; D021382:Protein Sorting Signals; D013329:Structure-Activity Relationship; D001618:beta-Lactamases / Q000378:metabolism*
D004274:DNA, Recombinant; D010455:Peptides; D011498:Protein Precursors; D021382:Protein Sorting Signals; D005914:Globins; D001618:beta-Lactamases