Novel low molecular weight organic compounds have been identified in thermophilic and hyperthermophilic Archaea and Bacteria that may play a role in osmotic and/or thermal adjustment (1). We used nuclear magnetic resonance spectroscopy to describe several novel solutes, namely, mannosylglycerate, mannosylglyceramide, di-2-O-b-mannosyl-di-myo-inositol-1,1'(3,3')-phosphate, di-myo-inositol-1,3'-phosphate, di-glycerolphosphate and b-galactopyranosyl-5-hydroxylysine (2-7). Thus far, we focused our attention on two of these solutes: mannosylglycerate and di-glycerolphosphate. Mannosylglycerate was identified and characterized in thermophilic bacteria, such as Rhodothermus marinus (3). Later this solute was detected in the hyperthermophilic archaea Pyrococcus furiosus and P. woesei (4), in Methanothermus sociabilis and M. fervidus (5), and in several organisms of the genus Thermococcus (6). In contrast to the wide distribution of mannosylglycerate, di-glycerolphosphate so far has been detected only in Archaeoglobus fulgidus (5). The fact that these solutes increase in concentration as the growth temperature and the salinity of the medium are raised, indicated that they could have a specific function in the thermostabilization of enzymes and other cellular components. The stabilizing effect of mannosylglycerate against heating or freeze-drying was, in fact, recently demonstrated on several enzymes from mesophilic, thermophilic and hyperthermophilic sources (8); results on the thermostabilizing effect of synthetically prepared di-glycerolphosphate will be presented as well. The effectiveness of these new solutes on the stabilization of model enzymes (lactate dehydrogenase, alcohol dehydrogenase, glucose oxidase, ribonuclease) as compared to that of classical stabilizers will be shown. These data disclose the potential of these compounds as enzyme stabilizers in biotechnological applications.

[1] M. S. da Costa, H. Santos and E. A. Galinski. An overview of the role and diversity of compatible solutes. Adv. Bioch. Eng. Biotechnol., 61, 117-153 (1998).

[2] L. O. Martins, L. S. Carreto, M. S. da Costa and H. Santos. New compatible solutes related to di-myo-inositol-phosphate in members of the order Thermotogales. J. Bacteriol., 178, 5644-5651 (1996).

[3] O. C. Nunes, C. M. Manaia, M. S. da Costa and H. Santos. Compatible solutes in thermophilic bacteria Rhodothermus marinus and "Thermus thermophilus". Appl. Environ. Microbiol., 61, 2351-2357 (1995).

[4] L. O. Martins and H. Santos. Accumulation of mannosylglycerate and di-myo-inositol-phosphate by Pyrococcus furiosus in response to salinity and temperature. Appl. Environ. Microbiol., 61, 3299-3303 (1995).

[5] L. O. Martins, R. Huber, H. Huber, K. O. Stetter, M. S. da Costa and H. Santos. Organic solutes in hyperthermophilic Archaea. Appl. Environ. Microbiol., 63, 896-902 (1997).

[6] P. Lamosa, L. O. Martins, M. S. da Costa and H. Santos. Effect of temperature, salinity, and the medium composition on compatible solute accumulation by Thermococcus spp. Appl. Environ. Microbiol. 64, 3591-98 (1998).

[7] Silva, Z., Borges, N., Martins, L. O., Wait, R., Da Costa, M. S., and Santos, H. (1999) Extremophiles 3, 163-172.

[8] A. Ramos, N. D. H. Raven, R. J. Sharp, S. Bartolucci, M. Rossi, R. Cannio, J. Lebbink, J. van der Oost, W. M. de Vos and H. Santos. Stabilization of enzymes against thermal stress and freeze-drying by mannosylglycerate. Appl. Env. Microbiol., 63, 4020-4025 (1997).