Measurement of protein phosphatase activity in biological samples using synthetic phosphopeptides.
A method has been developed for measuring specific protein phosphatase activity in biological samples using synthetic, phospho-Kemptide and phospho-GS-peptide. This method uses ion-exchange chromatography to determine phosphatase activity by quantifying the release of [32P]phosphate directly. The method was used to measure phosphatase activity of rat kidney, adrenals, heart, and liver cytosol and the activity of purified alkaline phosphatases, protein phosphatase 1, and protein phosphatase 2A. Ion-exchange chromatography was also used for the preparation of the radiolabeled phosphopeptide substrates. This method results in high recovery and specific activity of the labeled peptides. These techniques should be useful in isolating and characterizing specific protein phosphatases found in cells.
Nemani
R
R
Geriatric Research, Education, and Clinical Center, VA Medical Center, St. Louis, MO 63125.
Wongsurawat
N
N
Armbrecht
H J
HJ
D000818:Animals; D001681:Biological Assay; D002852:Chromatography, Ion Exchange; D008297:Male; D010748:Phosphopeptides; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2; D051381:Rats; D011916:Rats, Inbred F344; D013997:Time Factors
D010748:Phosphopeptides; D010761:Phosphorus Radioisotopes; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2