Age-dependent changes in extracellular proteins, aminopeptidase and proteinase activities in Frankia isolate BR.
To investigate protein secretion by the nitrogen-fixing actinomycete Frankia isolate BR, we designed a rapid DEAE adsorption, salt elution and Biogel P6DG desalination method to concentrate protein from the growth medium. Secreted proteins reached a maximum concentration (5.6 gm l-1) in the medium at growth arrest. Analysis by SDS-PAGE detected up to 63 extracellular polypeptides when Frankia cells were grown under stirred conditions in BAP medium supplemented with phosphatidylcholine and MES buffer and 65 proteins in stirred BAP media alone. The pattern of extracellular polypeptides changed during growth. Several extracellular proteolytic activities were detected and compared with intracellular ones. The substrate specificity of the extracellular and intracellular aminopeptidase activities were the same. Also, the electrophoretic migration patterns of secreted and intracellular aminopeptidases could not be distinguished. Secretion of the proline-specific aminopeptidase FAP proteinase (PF) were secreted: 10 had the same electrophoretic mobility as their intracellular counterparts after SDS-gelatine-PAGE while five (PF - 39.5, PF - 38.5, PF - 36.5, PF - 25.5 and PF - 20.5 kDa) had a different electrophoretic mobility and, therefore, appeared to be exclusively extracellular. At least seven extracellular proteinases appeared to increase coordinately in activity shortly before growth arrest.
Müller
A
A
Laboratoire d'Enzymologie, CNRS, 91198 Gif-sur-Yvette, France.
Benoist
P
P
Diem
H G
HG
Schwencke
J
J
D000626:Aminopeptidases / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology; D003470:Culture Media; D004586:Electrophoresis; D010450:Endopeptidases / Q000378:metabolism*; D040161:Frankia / Q000254:growth & development / Q000502:physiology*; D013997:Time Factors
D001426:Bacterial Proteins; D003470:Culture Media; D010450:Endopeptidases; D000626:Aminopeptidases