Ciulla R, Clougherty C, Belay N, Krishnan S, Zhou C, Byrd D, Roberts M F
Merkert Chemistry Center, Boston College, Chestnut Hill, Massachusetts 02167-3860.
J Bacteriol. 1994 Jun;176(11):3177-87. doi: 10.1128/jb.176.11.3177-3187.1994.
Methanobacterium thermoautotrophicum delta H and Marburg were adapted to grow in medium containing up to 0.65 M NaCl. From 0.01 to 0.5 M NaCl, there was a lag before cell growth which increased with increasing external NaCl. The effect of NaCl on methane production was not significant once the cells began to grow. Intracellular solutes were monitored by nuclear magnetic resonance (NMR) spectroscopy as a function of osmotic stress. In the delta H strain, the major intracellular small organic solutes, cyclic-2,3-diphosphoglycerate and glutamate, increased at most twofold between 0.01 and 0.4 M NaCl and decreased when the external NaCl was 0.5 M. M. thermoautotrophicum Marburg similarly showed a decrease in solute (cyclic-2,3-diphosphoglycerate, 1,3,4,6-tetracarboxyhexane, and L-alpha-glutamate) concentrations for cells grown in medium containing > 0.5 M NaCl. At 0.65 M NaCl, a new organic solute, which was visible in only trace amounts at the lower NaCl concentrations, became the dominant solute. Intracellular potassium in the delta H strain, detected by atomic absorption and 39K NMR, was roughly constant between 0.01 and 0.4 M and then decreased as the external NaCl increased further. The high intracellular K+ was balanced by the negative charges of the organic osmolytes. At the higher external salt concentrations, it is suggested that Na+ and possibly Cl- ions are internalized to provide osmotic balance. A striking difference of strain Marburg from strain delta H was that yeast extract facilitated growth in high-NaCl-containing medium. The yeast extract supplied only trace NMR-detectable solutes (e.g., betaine) but had a large effect on endogenous glutamate levels, which were significantly decreased. Exogenous choline and glycine, instead of yeast extract, also aided growth in NaCl-containing media. Both solutes were internalized with the choline converted to betaine; the contribution to osmotic balance of these species was 20 to 25% of the total small-molecule pool. These results indicate that M. thermoautotrophicum shows little changes in its internal solutes over a wide range of external NaCl. Furthermore, they illustrate the considerable differences in physiology in the delta H and Marburg strains of this organism.
嗜热自养甲烷杆菌δH菌株和马尔堡菌株被驯化以在含有高达0.65M NaCl的培养基中生长。在0.01至0.5M NaCl范围内，细胞生长前存在一个滞后期，且该滞后期随外部NaCl浓度的增加而延长。一旦细胞开始生长，NaCl对甲烷产生的影响并不显著。通过核磁共振（NMR）光谱监测细胞内溶质随渗透压应激的变化。在δH菌株中，主要的细胞内小分子有机溶质，环-2,3-二磷酸甘油酸和谷氨酸，在0.01至0.4M NaCl之间最多增加两倍，而当外部NaCl为0.5M时则减少。嗜热自养甲烷杆菌马尔堡菌株在含有>0.5M NaCl的培养基中生长的细胞，其溶质（环-2,3-二磷酸甘油酸、1,3,4,6-四羧基己烷和L-α-谷氨酸）浓度同样降低。在0.65M NaCl时，一种新的有机溶质成为主要溶质，该溶质在较低NaCl浓度下仅以痕量存在。通过原子吸收和³⁹K NMR检测到，δH菌株中的细胞内钾在0.01至0.4M之间大致恒定，然后随着外部NaCl进一步增加而降低。高细胞内K⁺由有机渗透物的负电荷平衡。在较高的外部盐浓度下，推测Na⁺以及可能的Cl⁻离子被内化以提供渗透平衡。马尔堡菌株与δH菌株的一个显著差异是酵母提取物促进了在含高NaCl培养基中的生长。酵母提取物仅提供痕量的可通过NMR检测到的溶质（例如甜菜碱），但对内源性谷氨酸水平有很大影响，使其显著降低。外源胆碱和甘氨酸而非酵母提取物也有助于在含NaCl培养基中的生长。两种溶质都被内化，胆碱转化为甜菜碱；这些物质对渗透平衡的贡献占小分子总量的20%至25%。这些结果表明，嗜热自养甲烷杆菌在广泛的外部NaCl范围内其细胞内溶质变化很小。此外，它们还说明了该生物体的δH菌株和马尔堡菌株在生理学上存在相当大的差异。
