[Stability of glucose 6-phosphate dehydrogenase complexed with its substrate and/or cofactor in aqueous and micellar environment].
Inactivation of glucose 6-phosphate dehydrogenase (G6PDH) complexed with its substrate, glucose 6-phosphate (GP), and/or cofactor, NADP+, has been studied within the range 20-40 degrees C in three media: (a) 0.04 M NaOH-glycine buffer (pH 9.1); (b) Aerosol OT (AOT) reversed micelles in octane; and (c) Triton X-100 micelles in octane supplemented with 10% hexanol. The enzyme inactivation was characterized quantitatively by first order rate constants, kin (s-1). In the case of G6PDH-NADP+ complexes, the values of kin were independent of the initial concentrations of G6PDH, either in aqueous medium or AOT micelles. The values of kin for the complex G6PDH-GP were inversely related to the initial concentration of the enzyme, in both aqueous and micellar media. When inactivation of both complexes were studied in AOT micelles, minimum values of kin corresponded to the degree of hydration W0 = 16.7; at W0 > 16.7 and W0 < 16.7, kin increased. Within the range 20-40 degrees C, the values of kin measured for both complexes in aqueous medium were significantly lower than those measured in AOT micelles. Temperature dependences of kin were characterized by inflections in Arrhenius plots, which corresponded, depending on the medium, to certain temperatures from 33.6 degrees C to 40 degrees C. In all media studied, NADP+ complexes of the enzyme exhibited higher stability than their GP counterparts. The parameters of G6PDH and G6PDH-NADP+ melting, measured by differential scanning microcalorimetry (maximum temperature and half-width of the transition, enthalpy of denaturation, and van't Hoff enthalpy), provided unequivocal evidence of the higher stability of the complex as compared to that of the enzyme. In addition, this approach demonstrated that G6PDH undergoes destabilization in AOT micelles.
Puchkaev
A V
AV
Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, 5/2 Kuprevich St., Minsk, 220141 Belarus.
Vlasov
A P
AP
Metelitsa
D I
DI
D002021:Buffers; D004143:Dioctyl Sulfosuccinic Acid; D004795:Enzyme Stability; D019298:Glucose-6-Phosphate / Q000737:chemistry / Q000378:metabolism; D005954:Glucosephosphate Dehydrogenase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008823:Micelles; D009249:NADP / Q000737:chemistry / Q000378:metabolism; D009795:Octanes; D017830:Octoxynol; D012997:Solvents; D013696:Temperature
D002021:Buffers; D008823:Micelles; D009795:Octanes; D012997:Solvents; D004143:Dioctyl Sulfosuccinic Acid; D009249:NADP; D019298:Glucose-6-Phosphate; D017830:Octoxynol; D005954:Glucosephosphate Dehydrogenase; C026728:octane