Membrane impermeable biotinylation reagent sulfo nhs ss biotin

Parental COS-7 cells and HEK293 cells were purchased from ATCC (Manassas, VA, USA). Hydroxychloroquine Sulfate and Chloroquine Disulfate were purchased from Sigma-Aldrich (St. Louis, MO, USA). Membrane-impermeable biotinylation reagent sulfo-NHS-SS-biotin, streptavidin agarose, protein G-agarose beads, and horseradish peroxidase-conjugated anti-mouse antibodies were obtained from Pierce Biotechnology (Rockford, IL, USA). We purchased [³H]-labeled estrone sulfate from PerkinElmer (Waltham, MA, USA). Mouse anti-myc antibody (9E10) was purchased from Roche (Indianapolis, IN, USA). Mouse anti-E-cadherin antibody was purchased from Abcam (Cambridge, MA, USA). Mouse anti-ubiquitin antibody, mouse anti-β-actin antibody and normal mouse IgG were purchased from Santa Cruz Biotechnology (Dallas, TX, USA). A 20S proteasome assay kit was purchased from Sigma-Aldrich (St. Louis, MO, USA). Normal mouse lgG, mouse monoclonal anti-ubiquitin and mouse monoclonal anti-β-actin antibodies were purchased from Santa Cruz (Santa Cruz, CA, USA).

Parental COS-7 cells were purchased from ATCC (Manassas, VA). [³H]-labeled para-aminohippuric acid was purchased from PerkinElmer (Waltham, MA). Membrane-impermeable biotinylation reagent sulfo-NHS-SS-biotin, streptavidin agarose beads, protein G-agarose beads, and protease inhibitor cocktail were obtained from Pierce Biotechnology (Rockford, IL). Plasmid for wild type human Nedd4–2 was generously provided by Dr. Peter Snyder from University of Iowa (Iowa City, IA). Site-directed mutagenesis kit was purchased from Agilent Technologies (Santa Clara, CA). Mouse anti-myc and anti-β-actin antibodies were purchased from Roche (Indianapolis, IN). Mouse anti-E-cadherin antibody was purchased from Abcam (Cambridge, MA). Mouse anti-FLAG antibody and anti-FLAG M2 affinity beads were purchased from Sigma-Aldrich (St. Louis, MO). Rabbit anti-phospho-Ser/Thr antibody was obtained from Cell Signaling Technology (Danvers, MA). Phorbol 12-myristate 13-acetate (PMA), staurosporine (St), and all other reagents were purchased from Sigma-Aldrich (St. Louis, MO).