1. Biochemistry and Chemical Biology
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Kinetics of initiating polypeptide elongation in an IRES-dependent system

  1. Haibo Zhang
  2. Martin Y Ng
  3. Yuanwei Chen
  4. Barry S Cooperman  Is a corresponding author
  1. Spark Therapeutics, United States
  2. University of Pennsylvania, United States
  3. 1 Hacker Way, United States
Research Article
  • Cited 16
  • Views 1,254
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Cite this article as: eLife 2016;5:e13429 doi: 10.7554/eLife.13429

Abstract

The intergenic IRES of Cricket Paralysis Virus (CrPV-IRES) forms a tight complex with 80S ribosomes capable of initiating the cell-free synthesis of complete proteins in the absence of initiation factors. Such synthesis raises the question of what effect the necessary IRES dissociation from the tRNA binding sites, and ultimately from all of the ribosome, has on the rates of initial peptide elongation steps as nascent peptide is formed. Here we report the first results measuring rates of reaction for the initial cycles of IRES-dependent elongation. Our results demonstrate that 1) the first two cycles of elongation proceed much more slowly than subsequent cycles, 2) these reduced rates arise from slow pseudo-translocation and translocation steps, and 3) the retarding effect of ribosome-bound IRES on protein synthesis is largely overcome following translocation of tripeptidyl-tRNA. Our results also provide a straightforward approach to detailed mechanistic characterization of many aspects of eukaryotic polypeptide elongation.

Article and author information

Author details

  1. Haibo Zhang

    Spark Therapeutics, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Martin Y Ng

    Department of Chemistry, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Yuanwei Chen

    1 Hacker Way, Menlo Park, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Barry S Cooperman

    Department of Chemistry, University of Pennsylvania, Philadelphia, United States
    For correspondence
    cooprman@pobox.upenn.edu
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Alan G Hinnebusch, National Institute of Child Health and Human Development, United States

Publication history

  1. Received: December 1, 2015
  2. Accepted: June 1, 2016
  3. Accepted Manuscript published: June 2, 2016 (version 1)
  4. Version of Record published: July 27, 2016 (version 2)

Copyright

© 2016, Zhang et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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