Probing the leucyl/phenylalanyl trna protein transferase active site with trna substrate analogues

Aminoacyl-trna protein transferases post-translationally conjugate an amino acid from an aminoacyl-trna onto the n-terminus of a target polypeptide. the eubacterial aminoacyl-trna protein transferase,l/f transferase,utilizes both leucyl-trnaleu and phenylalanyl-trnaphe as substrates. x-ray crystal structures with substrate analogues,the minimal substrate phenylalanyl adenosine (ra-phe) and inhibitor puromycin,have been used to characterize trna recognition by l/f transferase. however analyses of these two x-ray crystal structures reveal significant differences in binding. through structural analyses,mutagenesis,and enzymatic activity assays,we rationalize and demonstrate that the substrate analogues bind to l/f transferase with similar binding affinities using a series of different interactions by the various chemical groups of the analogues. our data also demonstrates that enlarging the hydrophobic pocket of l/f transferase selectively enhances puromycin inhibition and may aid in the development of improved inhibitors for this class of enzymes. © 2014 bentham science publishers.