Inactivation of bovine kidney beta-N-acetyl-D-glucosaminidase by nonenzymatic glucosylation.
Evidence is presented that the incubation of beta-N-acetyl-D-glucosaminidase from bovine kidney with glucose leads to the covalent incorporation of the sugar into the enzyme protein. Concomitantly, the enzyme activity becomes markedly reduced depending on the time of incubation and the concentration of glucose employed. The separate investigation of the isoenzymes A and B, as obtained after DEAE-cellulose chromatography of the preparation, revealed that isoenzyme A had lost some 80% of its initial activity after 15 days at 37 degrees C at 44.4mM glucose, whereas isoenzyme B activity remained unchanged. The inactivation of A was associated (1) with a marked decrease in protein stain intensity after polyacrylamide gel and Cellogel electrophoresis and (2) with the formation of a small amount of apparent isoenzyme B, as indicated by the following observations: (a) appearance of enzyme activity at the position of isoenzyme B after DEAE-cellulose chromatography, (b) appearance of protein with enzyme activity in the region of authentic isoenzyme B after Cellogel electrophoresis, (c) increase in molecular mass from 130 kDa to 205 kDa, (d) similarity in heat-stability and Km for p-nitrophenyl N-acetyl-beta-glucosaminide between glucose-treated isoenzyme A and authentic isoenzyme B. As in diabetes mellitus the activity of beta-N-acetyl-D-glucosaminidase has been reported to be decreased in kidney and other tissues, the possibility that nonenzymatic glucosylation of the enzyme might occur in vivo is considered.
D000118:Acetylglucosaminidase / Q000037:antagonists & inhibitors*; D000818:Animals; D002417:Cattle; D004355:Drug Stability; D005947:Glucose / Q000494:pharmacology*; D006596:Hexosaminidases / Q000037:antagonists & inhibitors*; D007527:Isoenzymes / Q000037:antagonists & inhibitors; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D008970:Molecular Weight
D007527:Isoenzymes; D006596:Hexosaminidases; D000118:Acetylglucosaminidase; D005947:Glucose