sentence: Proteoliposomes were added to the ubiquitination reaction mix containing buffer U ( 20mM HEPES ( pH 7.4 ) , 2mM magnesium acetate , 150mM sodium chloride , 0.5mM TCEP ) , 0.2mg / ml BSA 0.23 μ M Uba1p , 2.1 μ M Ubc4p , 2.3 μ M Ubc7p , 2.2 μ M Cue1p , 122.5 μ M His-ubiquitin ( Boston Biochem , from yeast ) , 2.5mM ATP .