title: An unusual effect of NADP+ on the thermostability of the nonphosphorylating glyceraldehyde-3-phosphate dehydrogenase from Streptococcus mutans.
abstract: Adiabatic differential scanning calorimetry was used to investigate the effect of NADP+ on the irreversible thermal denaturation of the nonphosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPN) from Streptococcus mutans. The GAPN-NADP+ binary complex showed a strongly decreased thermal stability, with a difference of about 20 °C between the temperatures of the thermal transition peak maxima of the complex and the free protein. This finding was similar to the previously described thermal destabilization of GAPN upon binding of inorganic phosphate to the substrate binding site and can be interpreted as the shift of the equilibrium between 2 conformers of tetrameric GAPN upon addition of the coenzyme. Single amino acid substitution, known to abolish the NADP+ binding, cancelled the calorimetric effect of the coenzyme. GAPN thermal inactivation was considerably decelerated in the presence of NADP+ showing that the apparent change in stability of the active centre can be the opposite to that of the whole protein molecule. NADP+ could also reactivate the inactive GAPN* species, obtained by the heating of the apoenzyme below the thermal denaturation transition temperature. These effects may reflect a mechanism that provides GAPN the sufficient flexibility for the earlier observed profound active site reorganizations required during the catalytic cycle. The elevated thermal stability of the apoenzyme may, in turn, be important for maintaining a constant level of active GAPN--an enzyme that is known to be crucial for the effective supply of the reducing equivalents in S. mutans and its ability to grow under aerobic conditions.
authors.affiliation: a Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119992, Russia.
authors.forename: Denis
authors.initials: D
authors.lastname: Arutyunov
authors.forename: Elena
authors.initials: E
authors.lastname: Schmalhausen
authors.forename: Victor
authors.initials: V
authors.lastname: Orlov
authors.forename: Sophie
authors.initials: S
authors.lastname: Rahuel-Clermont
authors.forename: Natalia
authors.initials: N
authors.lastname: Nagradova
authors.forename: Guy
authors.initials: G
authors.lastname: Branlant
authors.forename: Vladimir
authors.initials: V
authors.lastname: Muronetz
mesh_terms: D019943:Amino Acid Substitution; D001665:Binding Sites; D002152:Calorimetry, Differential Scanning; D002384:Catalysis; D004795:Enzyme Stability; D024601:Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009249:NADP / Q000378:metabolism*; D010766:Phosphorylation; D011489:Protein Denaturation; D020836:Protein Structure, Quaternary; D013295:Streptococcus mutans / Q000201:enzymology*; D013696:Temperature
chemical_list: D009249:NADP; D024601:Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+)