Purification, characterization, and primary structure of Escherichia coli protease VII with specificity for paired basic residues: identity of protease VII and OmpT.
Escherichia coli cells were found to contain a novel outer membrane-associated protease, designated protease VII (K. Sugimura and N. Higashi, J. Bacteriol. 170:3650-3654, 1988). This enzyme was purified to homogeneity and exhibited an apparent molecular weight of 36,000 on sodium dodecyl sulfate gels and 180,000 on a TSK G-3000SW column in the presence of Triton X-100. It was capable of cleaving several peptides at the center of paired basic residues but not at single basic residues, implying that it is distinct from trypsinlike proteases. Protease VII was most active at pH 6.0 and was sensitive to a serine protease inhibitor, diisopropylfluorophosphate, and to the bivalent cations Zn2+, Cu2+, and Fe2+. The nucleotide sequence of a protease VII gene-carrying DNA fragment, which had been cloned by complementation analysis (K. Sugimura, Biochem. Biophys. Res. Commun. 153:753-759, 1988) was determined. It carried two putative promoter regions and a putative Shine-Dalgarno sequence in addition to the complete structural gene, which encoded pre-protease VII of 317 amino acid residues, with the N-terminal 20 residues being a signal peptide. By comparing their amino acid sequences, protease VII and OmpT, which specifically cleaves ferric enterobactin receptor protein, were found to be identical.
Sugimura
K
K
Suntory Institute for Biomedical Research, Osaka, Japan.
Nishihara
T
T
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011487:Protein Conformation; D015183:Restriction Mapping; D012697:Serine Endopeptidases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D012697:Serine Endopeptidases; C057840:omptin outer membrane protease