SPECIFIC DETERMINATION OF TYROSINE-PHOSPHORYLATED PROTEINS AND PEPTIDES BY DIFFERENTIAL IODINATION

A new method for the selective and quantitative determination of phosp hotyrosine residues is presented using a differential iodination techn ique, Characterization of tyrosine-phosphorylated proteins was perform ed in a biological system using human U937 myeloid leukemia cells. The method is based on the saturation of free iodine binding sites using non-radioactive iodine. Samples are then treated with alkaline phospha tase. New iodine binding sites in dephosphorylated tyrosines are subse quently radio-iodinated, resulting in specific labeling of tyrosine ph osphates, Separation is performed by RP-HPLC or sodium dodecyl sulfate -polyacrylamide gel electrophoresis. Radiolabeled proteins are then id entified using a radioactivity detector or autoradiography.