Autolysis of propionibacteria: detection of autolytic enzymes by renaturing SDS-PAGE and additional buffer studies.
Five strains of propionibacteria with 70-90% autolysis in sodium lactate broth (SLB) were studied by renaturing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Several lytic bands ranging in size between 25 and 143 kDa were detected by using propionibacteria cells or cell walls as substrate in the gel. Four Propionibacterium freudenreichii strains showed similar autolytic-enzyme profiles, consisting of two autolytic bands, one with molecular mass 162 kDa and one in the range 123-143 kDa. However, the Propionibacterium acidipropionici strain showed a completely different profile, consisting of 8 autolytic bands with molecular masses of 122, 97, 71, 55, 43, 39, 31, and 25 kDa. Lytic enzymes from P. freudenreichii INF-alpha, P. freudenreichii ISU P-59, P. freudenreichii ISU P-24, and P. freudenreichii ISU P-50 showed lytic activity against cells from all these four strains, but not against P. acidipropionici ATCC 4965. However, P. acidipropionici ATCC 4965 autolysed only its own cells. Effects of pH, temperature, and ions on autolytic activity were tested by renaturing SDS-PAGE and in buffer systems. Results from the SDS-PAGE electrophoresis showed optimal autolytic activity of P. acidipropionici ATCC 4965 at 37 degrees C and in the pH range 7 to 8.5 and of P. freudenreichii ISU P-59 at 20 degrees C and in the pH range 5 to 7. The autolytic activity of P. acidipropionici ATCC 4965 was extremely heat stable (100 degrees C, 2 h), in contrast to the lytic activity of P. freudenreichii ISU P-59, which was heat labile. The autolytic activities of P. acidipropionici ATCC 4965 were inhibited by divalent cations, however, the lytic activities of P. freudenreichii ISU P-59 were activated by Mn(2+), Ca(2+), and Co(2+). In buffer, optimum autolysis of P. acidipropionici ATCC 4965 was observed at pH 8.5 and at 40 degrees C. P. freudenreichii ISU P-59 showed optimum autolysis in buffer at pH 7.5 and at 30 degrees C.
Ostlie
Hilde M
HM
Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N-1432 As, Norway. hilde.ostlie@umb.no
Vegarud
Gerd
G
Langsrud
Thor
T
D001433:Bacteriolysis / Q000187:drug effects / Q000502:physiology*; D002611:Cheese / Q000382:microbiology*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005516:Food Microbiology; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D011424:Propionibacterium / Q000201:enzymology*; D013696:Temperature