Fig. 1.
Fig. 1. Demonstration of the specificity of the GST-PAK1-glutathione-Sepharose 4B-beads for the GTP forms of rac and cdc42. / (A) Anti-rac immunoblot showing the effect of 100 μmol/L GDP, GTP, GDPβS, or GTPγS on the amount of rac collected from a 100 000gsupernatant (high-speed supernatant) of detergent-lysed human platelets. (B) Anti-cdc42 immunoblot showing the amount of cdc42 trapped after loading high-speed supernatant with 100 μmol/L GDP, GTP, GDPβS, or GTPγS. Rac or cdc42 remaining in the supernatant (S) was separated from that which bound to beads by centrifugation at 10 000g for 2 minutes. The bead pellets (P) were washed with excess volume of TBS-Tween 20 buffer (pH 7.4) and denatured with SDS-sample buffer, bound protein displayed by 12% PAGE, and then were transferred to PVDF membrane for immunoblotting.

Demonstration of the specificity of the GST-PAK1-glutathione-Sepharose 4B-beads for the GTP forms of rac and cdc42.

(A) Anti-rac immunoblot showing the effect of 100 μmol/L GDP, GTP, GDPβS, or GTPγS on the amount of rac collected from a 100 000gsupernatant (high-speed supernatant) of detergent-lysed human platelets. (B) Anti-cdc42 immunoblot showing the amount of cdc42 trapped after loading high-speed supernatant with 100 μmol/L GDP, GTP, GDPβS, or GTPγS. Rac or cdc42 remaining in the supernatant (S) was separated from that which bound to beads by centrifugation at 10 000g for 2 minutes. The bead pellets (P) were washed with excess volume of TBS-Tween 20 buffer (pH 7.4) and denatured with SDS-sample buffer, bound protein displayed by 12% PAGE, and then were transferred to PVDF membrane for immunoblotting.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal