Thermostabilization of proteins by diglycerol phosphate, a new compatible solute from the hyperthermophile Archaeoglobus fulgidus

Diglycerol phosphate accumulates under salt stress in the archaeon Archaeog lobus fulgidus (L. O. Martins, R. Huber, H. Huber, K. O. Stetter, M. S. da Costa, and H. Santos, Appl. Environ. Microbiol, 63:896-902, 1997), This sol ute was purified after extraction from the cell biomass. In addition, the o ptically active and the optically inactive (racemic) forms of the compound were synthesized, and the ability of the solute to act as a protecting agen t against heating was tested on several proteins derived from mesophilic or hyperthermophilic sources. Diglycerol phosphate exerted a considerable sta bilizing effect against heat inactivation of rabbit muscle lactate dehydrog enase, baker's yeast alcohol dehydrogenase, and Thermococcus litoralis glut amate dehydrogenase. Highly homologous and structurally well-characterized rubredoxins from Desulfovibrio gigas, Desulfovibrio desulfuricans (ATCC 277 74), and Clostridium pasteurianum were also examined for their thermal stab ilities in the presence or absence of diglycerol phosphate, glycerol, and i norganic phosphate. These proteins showed different intrinsic thermostabili ties, with half-lives in the range of 30 to 100 min. Diglycerol phosphate e xerted a strong protecting effect, with approximately a fourfold increase i n the half-lives for the loss of the visible spectra of D. gigas and C. pas teurianum rubredoxins. In contrast, the stability of D. desulfuricans rubre doxin was not affected. These different behaviors are discussed in the ligh t of the known structural features of rubredoxins. The data show that digly cerol phosphate is a potentially useful protein stabilizer in biotechnologi cal applications.