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Tly, collected, and centrifuged at 2000 rpm for 10 min before resuspension in cold lysis buffer containing 25 mM HEPES, pH 7.5, 150 mM NaCl, 1 NP-40, 10 mM MgCl2, 1 mM EDTA, and 10 glycerol (Upstate, Lake Placid, NY) and supplemented with protease inhibitor cocktail and phosphatase inhibitor cocktails 1 and 2 (1:100) (Sigma, St. Louis, MO). After incubation on ice for 30 minutes and subsequent c