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.
Full text
PDF
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Davidson V. L., Wu J., Miller B., Jones L. H. Factors affecting the stability of methanol dehydrogenase from Paracoccus denitrificans. FEMS Microbiol Lett. 1992 Jul 1;73(1-2):53–58. doi: 10.1016/0378-1097(92)90582-9. [DOI] [PubMed] [Google Scholar]
- Geiger O., Görisch H. Reversible thermal inactivation of the quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus. Ca2+ ions are necessary for re-activation. Biochem J. 1989 Jul 15;261(2):415–421. doi: 10.1042/bj2610415. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Harris T. K., Davidson V. L. A new kinetic model for the steady-state reactions of the quinoprotein methanol dehydrogenase from Paracoccus denitrificans. Biochemistry. 1993 Apr 27;32(16):4362–4368. doi: 10.1021/bi00067a028. [DOI] [PubMed] [Google Scholar]
- Harris T. K., Davidson V. L. Replacement of enzyme-bound calcium with strontium alters the kinetic properties of methanol dehydrogenase. Biochem J. 1994 May 15;300(Pt 1):175–182. doi: 10.1042/bj3000175. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nunn D. N., Day D., Anthony C. The second subunit of methanol dehydrogenase of Methylobacterium extorquens AM1. Biochem J. 1989 Jun 15;260(3):857–862. doi: 10.1042/bj2600857. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Richardson I. W., Anthony C. Characterization of mutant forms of the quinoprotein methanol dehydrogenase lacking an essential calcium ion. Biochem J. 1992 Nov 1;287(Pt 3):709–715. doi: 10.1042/bj2870709. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Salisbury S. A., Forrest H. S., Cruse W. B., Kennard O. A novel coenzyme from bacterial primary alcohol dehydrogenases. Nature. 1979 Aug 30;280(5725):843–844. doi: 10.1038/280843a0. [DOI] [PubMed] [Google Scholar]
- White S., Boyd G., Mathews F. S., Xia Z. X., Dai W. W., Zhang Y. F., Davidson V. L. The active site structure of the calcium-containing quinoprotein methanol dehydrogenase. Biochemistry. 1993 Dec 7;32(48):12955–12958. doi: 10.1021/bi00211a002. [DOI] [PubMed] [Google Scholar]