Enzymatic modification of proteins. 3. Purification and properties of a leucyl, phenylalanyl transfer ribonucleic acid protein transferase from Escherichia coli.

A leucyl, phenylalanyl transfer RNA-protein transferase has been purified nearly 300-fold from the 105,000 X g supernatant fraction of Escherichia coIi. The procedure consists of extraction of the precipitate formed with protamine sulfate, gel filtration on Sephadex G-100 and Sephadex G-75, and chromatography on diethylaminoethyl cellulose. The enzyme preparation specifically catalyzes the transfer of leucine and phenylalanine from tRNA to acceptor proteins. The transfer reaction requires the presence of monovalent cation, and maximal rates are obtained with 0.15 M KCl. The reaction is inhibited by divalent cations and by puromycin, but not by chloramphenicol. The transfer of both amino acids appears to be catalyzed by a single enzyme since the two activities are present in a constant ratio over the entire range of purification and show identical kinetics of thermal inactivation. When bovine serum albumin is employed as the acceptor protein, both amino acids are attached to it in a way that retains the free oc-amino group.