Protein phosphatase assay suitable for acid-labile substrates.
A sensitive protein phosphatase assay suitable for acid-labile substrates has been developed. The assay involves incubation of the phosphatase with a 32P-labeled phosphoprotein substrate, quenching the reaction, and then filtering through an ultrafilter that retains substrate and allows the released phosphate to flow through. The released phosphate is counted. The use of a slightly alkaline quenching solution makes the assay suitable for all phosphoamino acids that are stable near physiological pH, including the acid-labile phosphoamino acids. The process is made very convenient by the use of disposable microcon-3 ultrafiltration devices. Radioactive waste is limited to reaction tubes, the microcon-3s, pipette tips, and the scintillation vials and fluid.
Kim
Y
Y
Department of Biological Chemistry, School of Medicine, University of California, Davis 95616.
Matthews
H R
HR
D000143:Acids; D010749:Phosphoprotein Phosphatases / Q000032:analysis*; D010750:Phosphoproteins / Q000737:chemistry; D012680:Sensitivity and Specificity; D014462:Ultrafiltration
D000143:Acids; D010750:Phosphoproteins; D010749:Phosphoprotein Phosphatases