Phosphorylation of casein by human erythrocyte membrane-bound protein kinase: competition of casein with endogenous substrates.
The possibility that spectrin and band-3 protein are phosphorylated by the same membrane-bound protein kinase was investigated by adding casein to unsealed erythrocyte ghosts and examing competition of the three proteins for phosphorylation. The extent of spectrin and band-3 protein phosphorylation was reduced by up to approximately 55%. This indicated that casein was competing with these endogenous substrates for phosphorylation and was most probably phosphorylated by the same protein kinase(s). Furthermore, the extent of inhibition of the phosphorylation of the two endogenous substrates was indistinguishable over the range of casein concentrations tested (0.1 to 5 mg/ml). This indicates that spectrin and band-3 protein may be phosphorylated by the same protein kinase. In contrast, casein was found to have no effect on the cAMP-dependent phosphorylation of band 4.5. This result indicates that casein only competes with the endogenous proteins phosphorylated by the cAMP-independent protein kinase(s). The extent of reduction of endogenous substrate phosphorylation in the presence of casein was found to be constant over incubation periods of 1 to 15 min, indicating that this reduction was not due to consumption of ATP. Since the spectrin and band-3 protein phosphorylations were specifically and identically reduced by casein and these reductions were not due to the ATP consumption or to a general alteration of the membrane, we conclude that the two substrates are likely phosphorylated by one kinase which also phosphorylates casein.
D002364:Caseins / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology* / Q000378:metabolism; D000242:Cyclic AMP / Q000502:physiology; D004912:Erythrocytes / Q000201:enzymology* / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D013049:Spectrin / Q000378:metabolism
D002364:Caseins; D008565:Membrane Proteins; D013049:Spectrin; D000242:Cyclic AMP; D011494:Protein Kinases