Response of intracellular carbohydrates to a NaCl shock in Rhizobium leguminosarum biovar trifolii TA-1 and Rhizobium meliloti SU-47
Breedveld, M.W. | Dijkema, C. | Zevenhuizen, L.P.T.M. | Zehnder, A.J.B.
The dynamic response of cellular carbohydrates to a NaCl shock in Rhizobium leguminosarum biovar trifolii TA-1 (0.25 M-NaCl) and Rhizobium meliloti SU-47 (0.4 M-NaCl) grown in NaCl-free medium was investigated in nongrowing cell cultures and in cell suspensions, using in vivo NMR. After transferring cells grown in a NaCl-free medium to a glutamic-acid-free medium containing mannitol and NaCl, both strains immediately responded to the increased osmotic pressure by augmenting the cellular trehalose content of the cell. Without mannitol in the medium trehalose synthesis was slower, but clearly detectable. Its synthesis paralleled the breakdown of the reserve materials glycogen and poly-beta-hydroxybutyric acid (PHB). NMR experiments with 25-fold-concentrated cell suspensions using 13C1-mannitol as substrate revealed that 15-20% of the trehalose synthesized was derived from mannitol, but 80-85% was from other sources. Trehalose was mainly formed from the internal pool of glycogen and/or PHB, whether mannitol was present or not, and reached 135 and 280 micrograms (mg cell protein)-1 in the strains TA-1 and SU-47, respectively. At low osmolarity, intracellular trehalose was metabolized by strains TA-1 and SU-47. Intracellularly accumulated phosphoglycerol-substituted and neutral cyclic (1,2)-beta-glucans of SU-47 cells grown in the absence of NaCl were neither degraded nor excreted after exposure to NaCl. Strain TA-1, which only makes neutral cyclic (1,2)-beta-glucans, continued to synthesize and excrete cyclic (1,2)-beta-glucans after exposure to NaCl. By using in vivo 31P-NMR, a sharp peak at 1.34 p.p.m. was present in cell suspensions of strain SU-47. This peak, representing glycerol-1-phosphate-substituted cyclic glucans, was absent in strain TA-1.
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