Thermoanaerobacter ethanolicus 39E secondary-alcohol dehydrogenase (2 o ADH) was optimally active near 90 o C displaying thermostability half-lives of 1.2 days, 1.7 h, 19 min, 9.0 min, and 1.3 min at 80 o C, 90 o C, 92 o C, 95 o C, and 99 o C, respectively. Enzyme activity loss upon heating (90-100 o C) was accompanied by precipitation, but the soluble enzyme remaining after partial inactivation retained complete activity. Enzyme thermoinactivation was modeled by a pseudo-first order rate equation suggesting that the rate determining step was unimolecular with respect to protein and thermoinactivation preceded aggregation. The apparent 2 o ADH melting temperature (T m ) occurred at ~115 o C, 20 o C higher than the temperature for maximal activity, suggesting that it is completely folded in its active temperature range. Thermodynamic calculations indicated that the active folded structure of the 2 o ADH is stabilized by a relatively small Gibbs energy ( G s t a b . = 110 kJ mol - 1 ). 2 o ADH catalytic activities at 37 o C to 75 o C, were 2-fold enhanced by guanidine hydrochloride (GuHCl) concentrations between 120 mM and 190 mM. These results demonstrate the extreme resistance of this thermophilic 2 o ADH to thermal or chemical denaturation; and suggest increased temperature or GuHCl levels seem to enhance protein fixability and activity.