title: Thermal stability of methanol dehydrogenase is altered by the replacement of enzyme-bound Ca2+ with Sr2+.
abstract: Methanol dehydrogenase (MEDH) possesses tightly bound Ca2+ in addition to its pyrroloquinoline quinone prosthetic group. Ca2+ was replaced with Sr2+ by growing the host bacterium, Paracoccus denitrificans, in media in which Ca2+ was replaced with Sr2+. At temperatures in the transition region for stability, the rate constants for inactivation of MEDH purified from these cells (Sr-MEDH) were 2-fold lower than those for MEDH. However, Arrhenius plots yielded an activation energy (Ea) of 699 kJ (167 kcal)/mol for MEDH compared with 640 kJ (153 kcal)/mol for Sr-MEDH. Further analysis by transition-state theory yielded values for the activation enthalpy (delta H*) and activation entropy (delta S*) of 696 kJ (166 kcal)/mol and 1.73 kJ (414 cal)/mol per K for MEDH and 637 kJ (152 kcal)/mol and 1.55 kJ (371 cal)/mol per K for Sr-MEDH. The higher rate of inactivation of MEDH than Sr-MEDH at higher temperatures is a consequence of a more favourable net gain in entropy. This positive entropy contribution increases at high temperatures, and reduces the more favourable stability obtained from the enthalpy contribution for the free energy (delta G*) of inactivation. The differences in these thermodynamic data are discussed in relation to the recently determined crystal structure of MEDH as well as 1H electron-nuclear double resonance studies of the influence of Sr2+ substitution on the structure of the pyrroloquinoline quinone-derived radical in MEDH.
authors.affiliation: Department of Biochemistry, University of Mississippi Medical Center, Jackson 39216-4505.
authors.forename: T K
authors.initials: TK
authors.lastname: Harris
authors.forename: V L
authors.initials: VL
authors.lastname: Davidson
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D004795:Enzyme Stability; D007700:Kinetics; D010231:Paracoccus denitrificans / Q000201:enzymology*; D011485:Protein Binding; D013053:Spectrophotometry; D013324:Strontium / Q000378:metabolism* / Q000494:pharmacology; D013816:Thermodynamics
chemical_list: D000429:Alcohol Oxidoreductases; C019641:alcohol dehydrogenase (acceptor); D002118:Calcium; D013324:Strontium