“…The same purification protocol was used for Rab1b, although the respective wash buffer was complemented with 0.2 mM PMSF, 0.01 mM GDP, cOmplete protease inhibitor cocktail tablets (Roche) for cell lysis (Rab1b wash buffer: 40 mM HEPES pH 8, 2 mM ß-ME, 1 mM MgCl 2 , 500 mM LiCl, 0.01 mM GDP, 20 mM imidazole; Rab1b elution buffer: 40 mM HEPES pH 8, 2 mM ß-ME, 1 mM MgCl 2 , 500 mM LiCl, 0.01 mM GDP, 300 mM imidazole). 85,87 The incorporation of met-ncbK and quantitative decaging to metK was confirmed using full-length ESI-LC MS (Phenomenex Jupiter TM C4 column (2 × 150 mm, 5 μm)) and carried out on an Agilent Technologies 1260 Infinity LC-MS system with a 6310 quadrupole spectrometer. Buffer A was 0.1% formic acid in water; buffer B was 0.1% formic acid in acetonitrile.…”