HALOTOLERANCE IN METHANOSARCINA SPP - ROLE OF N-EPSILON-ACETYL-BETA-LYSINE, ALPHA-GLUTAMATE, GLYCINE BETAINE, AND K+ AS COMPATIBLE SOLUTES FOR OSMOTIC ADAPTATION

The methanogenic Archaea, like the Bacteria and Eucarya, possess sever al osmoregulatory strategies that enable them to adapt to osmotic chan ges in their environment. The physiological responses of Methanosarcin a species to different osmotic pressures were studied in extracellular osmolalities ranging from 0.3 to 2.0 osmol/kg. Regardless of the isol ation source, the maximum rate of growth for species from freshwater, sewage, and marine sources occurred in extracellular osmolalities betw een 0.62 and 1.0 osmol/kg and decreased to minimal detectable growth a s the solute concentration approached 2.0 osmol/kg, The steady-state w ater-accessible volume of Methanosarcina thermophila showed a dispropo rtionate decrease of 30% between 0.3 and 0.6 osmol/kg and then a linea r decrease of 22% as the solute concentration in the media increased f rom 0.6 to 2.0 osmol/kg. The total intracellular K+ ion concentration in M. thermophila increased from 0.12 to 0.5 mol/kg as the medium osmo lality was raised from 0.3 to 1.0 osmol/kg and then remained above 0.4 mol/kg as extracellular osmolality was increased to 2.0 osmol/kg, Con current with K+ accumulation, M. thermophila synthesized and accumulat ed alpha-glutamate as the predominant intracellular osmoprotectant in media containing up to 1.0 osmol of solute per kg, At medium osmolalit ies greater than 1.0 osmol/kg, the alpha-glutamate concentration level ed off and the zwitterionic beta-amino acid N-epsilon-acetyl-beta-lysi ne was synthesized, accumulating to an intracellular concentration exc eeding 1.1 osmol/kg at an osmolality of 2.0 osmol/kg, When glycine bet aine was added to culture medium, it caused partial repression of de n ovo alpha-glutalnate and N-epsilon-acetyl-beta-lysine synthesis and wa s accumulated by the cell as the predominant compatible solute, The di stribution and concentration of compatible solutes in eight strains re presenting five Methanosarcina spp, were similar to those found in M. thermophila grown in extracellular osmolalities of 0.3 and 2.0 osmol/k g, Results of this study demonstrate that the mechanism of halotoleran ce in Methanosarcina spp, involves the regulation of K+, alpha-glutama te, N-epsilon-acetyl-beta-lysine, and glycine betaine accumulation in response to the osmotic effects of extracellular solute.