Crystal structures of leucyl/phenylalanyl‐tRNA‐protein transferase and its complex with an aminoacyl‐tRNA analog

Eubacterial leucyl/phenylalanyl‐tRNA protein transferase (L/F‐transferase), encoded by the aat gene, conjugates leucine or phenylalanine to the N‐terminal Arg or Lys residue of proteins, using Leu‐tRNALeu or Phe‐tRNAPhe as a substrate. The resulting N‐terminal Leu or Phe acts as a degradation signal for the ClpS‐ClpAP‐mediated N‐end rule protein degradation pathway. Here, we present the crystal structures of Escherichia coli L/F‐transferase and its complex with an aminoacyl‐tRNA analog, puromycin. The C‐terminal domain of L/F‐transferase consists of the GCN5‐related N‐acetyltransferase fold, commonly observed in the acetyltransferase superfamily. The p‐methoxybenzyl group of puromycin, corresponding to the side chain of Leu or Phe of Leu‐tRNALeu or Phe‐tRNAPhe, is accommodated in a highly hydrophobic pocket, with a shape and size suitable for hydrophobic amino‐acid residues lacking a branched β‐carbon, such as leucine and phenylalanine. Structure‐based mutagenesis of L/F‐transferase revealed its substrate specificity. Furthermore, we present a model of the L/F‐transferase complex with tRNA and substrate proteins bearing an N‐terminal Arg or Lys.

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