Fraction 5 fatty acid free bsa

α-minimal essential medium, fetal bovine serum and antibiotic/antimycotic solution were from Life Technologies. All other reagent-grade chemicals, including BSA and fatty acids, were obtained from Sigma–Aldrich unless otherwise stated. Wortmannin and okadaic acid were from Calbiochem-Merck. Fraction V fatty acid-free BSA was from Roche. Insulin was from R&D systems. Complete protein phosphatase inhibitor tablets were purchased from Roche Diagnostics. Phospho-Akt^Thr308, phospho-Akt^Ser473, phospho-ERK1/2^{Thr202/Tyr204}, phospho-GSK3^Ser21/9, phospho-CREB^Ser133, phospho-Src^Tyr416, anti-Akt and anti-ERK1/2 antibodies were from Cell Signaling Technology. Anti-GSK3 and anti-PTEN antibodies were from Santa Cruz Biotechnology. Anti-IR (insulin receptor), anti-p85, anti-IRS1 and anti-PP2A subunit C, demethylated were from Millipore. Anti-β actin and anti-GAPDH were from Sigma. Phospho-PP2A^Y307 was from Abcam.

α-MEM (α-minimal essential medium), DMEM (Dulbecco's modified Eagle's medium), EBSS (Earle's Balanced Salt Solution), Opti-Mem, FBS (fetal bovine serum), and antibiotic/antimycotic solution were from Life Technologies. All other reagent-grade chemicals, including BSA and fatty acids, were obtained from Sigma-Aldrich unless otherwise stated. Fraction V fatty acid-free BSA was from Roche. Complete protein phosphatase inhibitor tablets were purchased from Boehringer-Roche Diagnostics. Anti-Slc38a2 (SNAT2) (catalogue BMP081) was from MBL, anti-ubiquitin (catalogue Z0458) was from Dako, anti-K48 (catalogue ab140601), and anti-K63 (catalogue ab179434) linkage specific ubiquitin antibodies were from Abcam, anti-α-subunit Na/K-ATPase (catalogue a6F) was from DSHB, anti-β actin(catalogue A5060), anti-GAPDH (catalogue G9545), and anti-Flag (catalogue F3165) were from Sigma, anti-Nedd4.2 and pCMV5-ubiquitin-Flag vector were obtained from Division of Signal Transduction Therapy (University of Dundee). Peptide-N-glycosidase F (PNGase F) was purchased from New England Biolabs. Go TaqDNA polymerase, deoxyribonucleotide phosphates (dNTPs) and Moloney Murine Leukemia Virus (M-MLV) reverse transcriptase were purchased from Promega. All primers were synthesized by the Oligonucleotide Synthesis Service (University of Dundee). X-ray film was purchased from Konica Minolta (Tokyo, Japan).