Purification and properties of fMet-tRNAf deacylase from Escherichia coli

Abstract

A formylmethionyl-tRNA_f deacylase has been purified about 330-fold from a crude initiation factor preparation (1 M NH₄Cl ribosomal wash) from Escherichia coli Q13. The enzyme was nearly homogeneous and had an apparent molecular weight of 24 000. Rat liver methionyl-tRNA_f and E. coli methionyl-tRNA_m were not hydrolyzed significantly by the enzyme under standard conditions. Qβ RNA- and AUG(A)_n-directed polypeptide synthesis was inhibited by the enzyme. The inhibition was at the level of initiation of polypeptide synthesis. The enzymatic activity was inhibited by various factors necessary for polypeptide synthesis. The activity was inhibited more by NH₄Cl and spermidine than by Mg²⁺, GTP and ATP. The complex of formylmethionyl-tRNA_f, initiation factor 2 and GTP was resistant to enzymatic hydrolysis, and the resistance was enhanced by the addition of AUG and ribosomes to the above reaction mixture.