Switching osmolyte strategies: response of Methanococcus thermalithotrophicus to changes in external NaCl

Methanococcus thermolifhatrophicus, a thermophilic methanogenic archaeon, p roduces and accumulates beta -glutamate and L-alpha -glutamate as osmolytes when grown in media with < 1 M NaCl. When the organism is adapted to grow in > 1 M NaCl, a new zwitterionic solute, N-E-acetyl-beta -lysine, is synth esized and becomes the dominant osmolyte. Several techniques, including in vivo and in vitro NMR spectroscopy, HPLC analyses of ethanol extracts, and potassium atomic absorption, have been used to monitor the immediate respon se of M. thermolithotrophicus to osmotic stress. There is a temporal hierar chy in the response of intracellular osmoljites. Changes in intracellular K + occur within the first few minutes of altering the external NaCl. Upon hy poosmotic shock, Ki is released from the cell; relatively small changes occ ur in the organic osmolyte pool on a longer time scale. Upon hyperosmotic s hock, 1M. thermolithotrophicus immediately internalizes K+, far more than w ould be needed stoichiometrically to balance the new salt concentration. Th is is followed by a decrease to a new K+ concentration (over 10-15 min), at which point synthesis and accumulation of primarily L-alpha -glutamate occ ur. Once growth of the M, thermolithotrophicus culture begins, typically 30 -100 min after the hyperosmotic shock, the intracellular levels of organic anions decrease and the zwitterion (NE-acetyl-beta -lysine) begins to repre sent a larger fraction of the intracellular pool. The observation that NE-a cetyl-beta -lysine accumulation occurs in osmoadapted cells but not immedia tely after osmotic shock is consistent with the hypothesis that lysine 2,3- aminomutase, an enzyme involved in NE-acetyl-beta -lysine synthesis, is eit her not present at high levels or has low activity in cells grown and adapt ed to lower NaCl. That lysine aminomutase specific activity is 8-fold lower in protein extracts from cells adapted to low NaCl compared to those adapt ed to 1.4 M NaCl supports this hypothesis. (C) 2000 Elsevier Science B.V. A ll rights reserved.