Interconversion of Two Forms of Leucyl-tRNA Synthetase
Two forms of leucyl-tRNA synthetase were isolated from E. Coli. Both forms, named EI and EII, catalyse the formation of leucyladenylate and the ATP-pyrophosphate exchange reaction with identical Km for leucine and ATP. Each form binds specifically tRNAs acceptor of leucine but only EII is able to transfer the activated aminoacid onto tRNA. Moreover if EII is highly specific for leucine, EI catalyses pyrophosphate exchange also in the presence of valine, isoleucine and methionine. The titration of sulfhydryl groups shows that two groups are masked in EI. One SH group in EII is rapidly reacting with pMB and is essential to the transfer of leucine to tRNA. EII with one SH blocked resembles EI by its catalytic properties but remains specific for leucine and cannot be separated from native EII. The two forms EI and EII are interconvertible in the presence of cell extracts. It was found that two factors are involved In this process; F1 which is a protease and releases a peptide of 3 000 MW from EII, thereby converting EII into EI. This peptide which is the factor F2 can reassociate with EI leading to an enzyme with catalytic and Chromatographic properties of EII. The results indicate that EII is constituted from two subunit-like fragments which are in EI associated by low-energy bonds and in the EII also by two peptide bonds at the ends of peptide F2. The association of F2 with the two “subunits” is necessary for full EII activity. Low-energy bonds are sufficient to keep this association (EI−F1) and reconstituted enzyme from EI and F1 maintains a conformation and properties close to those of native EII.
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