STUDIES ON THE SUBCELLULAR-FRACTIONS OF AEROMONAS-SALMONICIDA GROWN AT DIFFERENT TEMPERATURES

Aeromonas salmonicida cells were separated into cytoplasmic and peripl asmic fractions using osmotic shock in medium of 20% sucrose-1 mM EDTA . Using this method, less than 1% of the cytoplasmic glutamate dehydro genase contaminated the periplasmic fraction. The inner and outer memb rane fractions were separated by differential solubilization using the detergent Sarkosyl. All four fractions, together with the extracellul ar proteins, showed distinctive profiles on SDS-polyacrylamide gel ele ctrophoresis. A prominent protein of M(r) 42 000, present in the outer membrane fraction, had the distinctive properties of being resistant to proteolysis, extractable using 0.1 m NaCl/2% SDS, but not readily d issociated from the outer membrane on incubation with 2% SDS for 30 mi n at 37-degrees-C. When A. salmonicida was grown at 32-degrees-C, ther e was considerable reduction in the secretion of the 70-kDa protease, amylase and deoxyribonuclease, but there was also extensive cell lysis , as evident from the release of glutamate dehydrogenase. The proteins present in the culture filtrate included some from other cell fractio ns.