title: Modulation of the epithelial sodium channel (ENaC) by bacterial metalloproteases and protease inhibitors.
abstract: The serralysin family of metalloproteases is associated with the virulence of multiple gram-negative human pathogens, including Pseudomonas aeruginosa and Serratia marcescens. The serralysin proteases share highly conserved catalytic domains and show evolutionary similarity to the mammalian matrix metalloproteases. Our previous studies demonstrated that alkaline protease (AP) from Pseudomonas aeruginosa is capable of activating the epithelial sodium channel (ENaC), leading to an increase in sodium absorption in airway epithelia. The serralysin proteases are often co-expressed with endogenous, intracellular or periplasmic inhibitors, which putatively protect the bacterium from unwanted or unregulated protease activities. To evaluate the potential use of these small protein inhibitors in regulating the serralysin induced activation of ENaC, proteases from Pseudomonas aeruginosa and Serratia marcescens were purified for characterization along with a high affinity inhibitor from Pseudomonas. Both proteases showed activity against in vitro substrates and could be blocked by near stoichiometric concentrations of the inhibitor. In addition, both proteases were capable of activating ENaC when added to the apical surfaces of multiple epithelial cells with similar slow activation kinetics. The high-affinity periplasmic inhibitor from Pseudomonas effectively blocked this activation. These data suggest that multiple metalloproteases are capable of activating ENaC. Further, the endogenous, periplasmic bacterial inhibitors may be useful for modulating the downstream effects of the serralysin virulence factors under physiological conditions.
authors: Department of Cell Biology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania, United States of America.
authors: Michael B
authors: MB
authors: Butterworth
authors: Department of Cell Biology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania, United States of America.
authors: Liang
authors: L
authors: Zhang
authors: Department of Cell Biology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania, United States of America.
authors: Xiaoning
authors: X
authors: Liu
authors: Department of Ophthalmology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania, United States of America.
authors: Robert M
authors: RM
authors: Shanks
authors: Department of Cell Biology, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania, United States of America.
authors: Patrick H
authors: PH
authors: Thibodeau
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D001980:Bronchi / Q000166:cytology; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D010450:Endopeptidases / Q000737:chemistry / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D006801:Humans; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D011480:Protease Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D017510:Protein Folding; D059748:Proteolysis / Q000187:drug effects; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D051381:Rats; D012706:Serratia marcescens / Q000201:enzymology
chemical_list: D001426:Bacterial Proteins; D053503:Epithelial Sodium Channels; D011480:Protease Inhibitors; D010450:Endopeptidases; D008666:Metalloendopeptidases; C095666:serralysin; C087822:alkaline protease; D002118:Calcium