Characteristics of the biologically active 35-kDa metalloprotease virulence factor from Listeria monocytogenes.
Listeria monocytogenes, a facultative intracellular pathogen, synthesizes an extracellular protease which is responsible for the maturation of phosphatidylcholine phospholipase C (lecithinase), a virulence factor involved in cell-to-cell spread. This work describes the environmental parameters necessary for increased production of mature, 35-kDa active protease in strains of L. monocytogenes, and its detection using polyclonal antibodies raised against Bacillus subtilis neutral protease. High performance liquid affinity chromatography was exploited to isolate the biologically active form of the mature protease, which was then subjected to biochemical characterization using casein as a substrate. The protease is a zinc-dependent metalloprotease which degrades casein over a wide range of temperatures and pH values. It can also degrade actin, the most abundant protein in many eukaryotic cells. The Listeria protease was shown to exhibit a high thermal stability and a relatively narrow substrate specificity. A three-dimensional model built on the basis of the homology with thermolysin was used to understand the structural basis of these characteristics.
Coffey
A
A
Laboratory of Food Microbiology, Wageningen Agricultural University, The Netherlands.
van den Burg
B
B
Veltman
R
R
Abee
T
T
D000199:Actins / Q000378:metabolism; D002364:Caseins / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006361:Heating; D008089:Listeria monocytogenes / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D008958:Models, Molecular; D011480:Protease Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity; D013820:Thermolysin / Q000737:chemistry; D014774:Virulence
D000199:Actins; D002364:Caseins; D011480:Protease Inhibitors; D008666:Metalloendopeptidases; D013820:Thermolysin