Cell-bound and secreted proteases of Serratia marcescens.
Exoprotease of Serratia marcescens ATCC 25419 is exceptional among members of the family Enterobacteriaceae in that it is secreted in large amounts by viable cells into the culture medium. Labeling of cells with radioactive amino acids revealed no intracellular protein that could be precipitated with antibodies raised against purified exoproteases. With substances known to interfere with the excretion of some proteins--tosyl-L-lysine chloromethyl ketone, phenethyl alcohol, procaine, and sodium azide--and with rifampin, an intracellular form (apparent molecular weight, 52,000) larger than the major exoform (molecular weight, 51,000) was identified. Moreover, the 52,000-molecular-weight form was the main protein in immunoprecipitates of a cysteine-auxotrophic mutant starved for cysteine. Beside the major exoform, protease I, two additional exoproteases, termed II and III, appeared in the medium of stationary cultures. They were precipitated by antibodies against protease I, were identical in the Ouchterlony double-diffusion assay, and exhibited only a small difference, if any at all, in the peptide pattern after partial hydrolysis with protease V8 of Staphylococcus aureus. The amino- and carboxy-terminal amino acid sequences of protease I and II were determined and found to be identical, NH2-Ala-Ala-Thr-Gly-Gly-Tyr-Asp-Ala-Val-Asp and Phe-Ile-Val-COOH, respectively. The microheterogeneity of the isolated exoforms revealed by anion-exchange chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis was also observed in samples pulse-labeled with radioactive amino acids. It remains to be determined whether the different protease forms are the result of processing (modification) reactions or whether they constitute isoenzymes encoded by very similar genes.
Schmitz
G
G
Braun
V
V
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002451:Cell Compartmentation; D003545:Cysteine / Q000378:metabolism; D000939:Epitopes; D010447:Peptide Hydrolases / Q000032:analysis / Q000276:immunology / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D012706:Serratia marcescens / Q000201:enzymology*
D000596:Amino Acids; D000939:Epitopes; D011498:Protein Precursors; D010447:Peptide Hydrolases; D003545:Cysteine