A comparative infrared spectroscopic study of glycoside hydrolases from extremophilic Archaea revealed different molecular mechanisms of adaptation to high temperatures

The identification of the determinants of protein thermal stabilization is often pursued by comparing enzymes from hyperthermophiles with their mesophilic counterparts while direct structural comparisons among proteins and enzymes from hyperthermophiles are rather uncommon. Here, oligomeric b-glycosidases from the hyperthermophilic archaea Sulfolobus solfataricus (Ssb-gly), Thermosphaera aggregans (Tabgly), and Pyrococcus furiosus (Pfb-gly), have been compared. Studies of FTIR spectroscopy and kinetics of thermal inactivation showed that the three enzymes had similar secondary structure composition, but Ssb-gly and Tab-gly (temperatures of melting 98.1 and 98.48C, respectively) were less stable than Pfb-gly, which maintained its secondary structure even at 99.58C. The thermal denaturation of Pfb-gly, followed in the presence of SDS, suggested that this enzyme is stabilized by hydrophobic interactions. A detailed inspection of the 3D-structures of these enzymes supported the experimental results: Ssb-gly and Tab-gly are stabilized by a combination of ion-pairs networks and intrasubunit S-S bridges while the increased stability of Pfb-gly resides in a more compact protein core. The different strategies of protein stabilization give experimental support to recent theories on thermophilic adaptation and suggest that different stabilization strategies could have been adopted among archaea.