Characterization of a cytotoxic factor in culture filtrates of Serratia marcescens.
Serratia marcescens culture filtrates have been reported to be cytotoxic to mammalian cells. Using biochemical and genetic approaches, we have identified a major source of this cytotoxic activity. Both heat and protease treatments abrogated the cytotoxicity of S. marcescens culture filtrates towards HeLa cells, suggesting the involvement of one or more protein factors. A screen for in vitro cytotoxic activity revealed that S. marcescens mutant strains that are deficient in production of a 56-kDa metalloprotease are significantly less cytotoxic to mammalian cells. Cytotoxicity was significantly reduced when culture filtrates prepared from wild-type strains were pretreated with either EDTA or 1,10-phenanthroline, which are potent inhibitors of the 56-kDa metalloprotease. Furthermore, cytotoxic activity was restored when the same culture filtrates were incubated with zinc divalent cations, which are essential for enzymatic activity of the 56-kDa metalloprotease. Finally, recombinant expression of the S. marcescens 56-kDa metalloprotease conferred a cytotoxic phenotype on the culture filtrates of a nonpathogenic Escherichia coli strain. Collectively, these data suggest that the 56-kDa metalloprotease contributes significantly to the in vitro cytotoxic activity commonly observed in S. marcescens culture filtrates.
Marty
Kent B
KB
Department of Biology and Biochemistry, University of Houston, Houston, Texas 77204-5001, USA.
Williams
Christopher L
CL
Guynn
Linda J
LJ
Benedik
Michael J
MJ
Blanke
Steven R
SR
D001427:Bacterial Toxins / Q000235:genetics / Q000494:pharmacology*; D003603:Cytotoxins / Q000235:genetics / Q000494:pharmacology*; D004926:Escherichia coli / Q000235:genetics / Q000472:pathogenicity; D006367:HeLa Cells; D006801:Humans; D008666:Metalloendopeptidases / Q000235:genetics / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D012706:Serratia marcescens / Q000472:pathogenicity*; D015032:Zinc
D001427:Bacterial Toxins; D003603:Cytotoxins; D011994:Recombinant Proteins; D008666:Metalloendopeptidases; C044194:microbial metalloproteinases; D015032:Zinc