American Society of Hematology

Figure 5.

$Figure 5. Phosphorylation of tubulin by FK2 immunoprecipitates. (A) Washed platelets were activated with convulxin and lysed in NP-40 buffer. Lysates were treated with FK2 antibody for 1 hour at 4°C followed by 1 hour with Protein A/G PLUS-agarose. The immunoprecipitates were assessed by an in vitro kinase assay in the presence of DMSO (• and —) or 2 μM PP2 (▵ and —) or 200 μM piceatannol (□ and –). The inset shows the blot from which these data were derived. (B) The FK2 fraction of convulxin-activated platelets (• and —) and the total Syk fraction (▵ and –) were assayed for their kinase activity as in Figure 4. The relative protein mass in each fraction was assessed from the immunoblots and activity calculated based on the relative mass. Since the mass of the total Syk was about 10-fold higher than the polyubiquitinated fraction, we also assayed the total Syk fraction after 10-fold dilution (□ and —).$

Phosphorylation of tubulin by FK2 immunoprecipitates. (A) Washed platelets were activated with convulxin and lysed in NP-40 buffer. Lysates were treated with FK2 antibody for 1 hour at 4°C followed by 1 hour with Protein A/G PLUS-agarose. The immunoprecipitates were assessed by an in vitro kinase assay in the presence of DMSO (• and —) or 2 μM PP2 (▵ and —) or 200 μM piceatannol (□ and –). The inset shows the blot from which these data were derived. (B) The FK2 fraction of convulxin-activated platelets (• and —) and the total Syk fraction (▵ and –) were assayed for their kinase activity as in Figure 4. The relative protein mass in each fraction was assessed from the immunoblots and activity calculated based on the relative mass. Since the mass of the total Syk was about 10-fold higher than the polyubiquitinated fraction, we also assayed the total Syk fraction after 10-fold dilution (□ and —).

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