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Protein & Peptide Letters

Editor-in-Chief

ISSN (Print): 0929-8665
ISSN (Online): 1875-5305

Probing the Leucyl/Phenylalanyl tRNA Protein Transferase Active Site with tRNA Substrate Analogues

Author(s): Angela Wai Shan Fung, H. Alexander Ebhardt, Kollappillil S. Krishnakumar, Jack Moore, Zhizhong Xu, Peter Strazewski and Richard P. Fahlman

Volume 21, Issue 7, 2014

Page: [603 - 614] Pages: 12

DOI: 10.2174/0929866521666140212110639

Price: $65

Abstract

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.

Keywords: Aminoacyl-tRNA protein transferase, L/F transferase, N-end rule, puromycin, quantitative mass spectrometry.

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