Psychrophiles, host of permanently cold habitats, display metabolic fluxes comparable to those exhibited by mesophilic organisms at moderate temperatures. These organisms have evolved by producing, among other peculiarities, cold-active enzymes that have the properties to cope with the reduction of chemical reaction rates induced by low temperatures. The emerging picture suggests that these enzymes display a high catalytic efficiency at low temperatures through an improved flexibility of the structural components involved in the catalytic cycle, whereas other protein regions, if not implicated in catalysis, may be even more rigid than their mesophilic counterparts. In return, the increased flexibility leads to a decreased stability of psychrophilic enzymes. In order to gain further advances in the analysis of the activity/flexibility/stability concept, psychrophilic, mesophilic, and thermophilic DNA ligases have been compared by three-dimensional-modeling studies, as well as regards their activity, surface hydrophobicity, structural permeability, conformational stabilities, and irreversible thermal unfolding. These data show that the cold-adapted DNA ligase is characterized by an increased activity at low and moderate temperatures, an overall destabilization of the molecular edifice, especially at the active site, and a high conformational flexibility. The opposite trend is observed in the mesophilic and thermophilic counterparts, the latter being characterized by a reduced low temperature activity, high stability and reduced flexibility. These results strongly suggest a complex relationship between activity, flexibility and stability. In addition, they also indicate that in coldadapted enzymes, the driving force for denaturation is a large entropy change.The temperature range in which biological activity has been detected extends from Ϫ20°C, the temperature recorded in the brine veins of Arctic or Antarctic sea ice (1), to 113°C, the temperature at which the archae Pyrolobus fumarii is still able to grow (2). Although numerous investigations have been carried out on thermophilic microorganisms and on their molecular components, especially enzymes, the efforts devoted to coldadapted microorganisms have been comparatively limited despite their tremendous biotechnological (1, 3-5) and fundamental (1, 6 -8) applications. Indeed, the biochemical and physiological bases of cold adaptation, which include, for example, regulation of gene expression by low temperatures, membrane adaptation in relation with the homeoviscosity concept, and the activity/stability relationships sustaining the catalytic efficiency of cold-adapted enzymes, are still poorly understood.In permanent cold habitats, low temperatures have constrained psychrophiles to develop among other peculiarities enzymatic tools allowing metabolic rates compatible to life that are close to those of temperate organisms. Thermal compensation in these enzymes is reached, in most cases, through a high catalytic efficiency at low and moderate temperatures (for revie...