Fig. 5. One-dimensional phosphopeptide mapping analysis of erythroid ankyrin isoforms. Ankyrin immunoprecipitates were prepared from the detergent soluble (lanes 1-3) and insoluble fractions (lanes 4-8) of control (lanes 5-8) or calyculin A treated erythroid cells (lanes 1-4). Some of the precipitates were prepared from cells labeled with ³²P-orthophosphate (lanes 3-5). The remaining precipitates, which were prepared from unlabeled cells, were phosphorylated in vitro by the coprecipitating kinase (lanes 2, 6 and 7) or by purified CK2 (lanes 1 and 8). In each instance, the precipitates were resolved on a 6% SDS polyacrylamide gel, and the gel was stained with GelCode Blue. The indicated ankyrin isoforms were excised from the gel, electroeluted, and the eluted proteins were digested with V8 endoproteinase. The resulting phosphopeptides were analyzed on 18% SDS polyacrylamide gels. The gels were dried, and ³²P-labeled peptides were detected by autoradiography. Molecular weight markers are indicated to the left of the figure.

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