RESUMO
Effects of alloxan-diabetes on kinetic attributes of Na(+)+K(+)-ATPase were examined in rat kidney, brain and erythrocyte membranes. The enzyme activity decreased significantly from 60-80% in the three membrane systems as a result of diabetic state. Kinetic analysis revealed that Km of the enzyme increased by 5- and 10-fold respectively in the kidney and brain membranes while registering a 50-60% decrease in Vmax. Ouabain binding studies revealed that in the kidney membranes the I50 value increased by 150-fold in diabetic animals with a significant decrease in number of ouabain molecules bound; at concentrations beyond 10(-7) M de-binding of ouabain occurred. For the brain membranes the I50 values for ouabain increased even more significantly (2000-fold increase) without any change in Hill coefficient for ouabain binding. Glycosylation studies revealed that its extent was highest for the brain and least for the kidney membranes which correlated to some extent with the I50 and Km values but not with Vmax. The results thus suggest that glycosylation in critical domains of the membrane and/or enzyme structure may play an important regulatory role. Physiological significance of these findings is discussed.