Sequence of the leader peptidase gene of Escherichia coli and the orientation of leader peptidase in the bacterial envelope.
The nucleotide sequence of the leader peptidase structural gene from Escherichia coli has been determined. The gene codes for a protein of 323 amino acid residues with a calculated Mr = 35,994, in agreement with the apparent molecular weight of leader peptidase (37,000) determined from gel electrophoresis in sodium dodecyl sulfate. In addition, the amino acid composition predicted from the DNA sequence matches that of the purified enzyme. Leader peptidase synthesized in a cell-free transcription-translation system and isolated from cell membranes have identical apparent molecular weights on sodium dodecyl sulfate-polyacrylamide gels. The presence of methionine and cysteine in the NH2-terminal tryptic peptide shows that no more than three amino acid residues have been removed from the primary translation product predicted from the DNA sequence. These results indicate that leader peptidase is synthesized and assembled into the membrane without proteolytic removal of a leader peptide. Leader peptidase is shown to be largely found in the inner membrane of overproducing strains. The majority of the polypeptide chain is exposed on the outer surface of the inner membrane. It is anchored by a membrane-spanning segment near the NH2 terminus. This orientation agrees with the functional orientation of leader peptidase observed in artificial membrane vesicles.
D000595:Amino Acid Sequence; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology / Q000648:ultrastructure; D004262:DNA Restriction Enzymes; D010450:Endopeptidases / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology*; D005796:Genes; D005798:Genes, Bacterial; D008565:Membrane Proteins; D010446:Peptide Fragments / Q000032:analysis; D012697:Serine Endopeptidases; D014357:Trypsin
D008565:Membrane Proteins; D010446:Peptide Fragments; D004262:DNA Restriction Enzymes; D010450:Endopeptidases; D012697:Serine Endopeptidases; D014357:Trypsin; C026515:type I signal peptidase