The ribosome can discriminate the chirality of amino acids within its peptidyl-transferase center

Michael T. Englander, Joshua L. Avins, Rachel C. Fleisher, Bo Liu, Philip R. Effraim, Jiangning Wang, Klaus Schulten, Thomas S. Leyh, Ruben L. Gonzalez, Virginia W. Cornish

Research output: Contribution to journalArticle

36 Scopus citations

Abstract

The cellular translational machinery (TM) synthesizes proteins using exclusively L- or achiral aminoacyl-tRNAs (aa-tRNAs), despite the presence of D-amino acids in nature and their ability to be aminoacylated onto tRNAs by aa-tRNA synthetases. The ubiquity of L-amino acids in proteins has led to the hypothesis that D-amino acids are not substrates for the TM. Supporting this view, protein engineering efforts to incorporate D-amino acids into proteins using the TM have thus far been unsuccessful. Nonetheless, a mechanistic understanding of why D-aa-tRNAs are poor substrates for the TM is lacking. To address this deficiency, we have systematically tested the translation activity of D-aa-tRNAs using a series of biochemical assays. We find that the TM can effectively, albeit slowly, accept D-aa-tRNAs into the ribosomal aa-tRNA binding (A) site, use the A-site D-aa-tRNA as a peptidyl-transfer acceptor, and translocate the resulting peptidyl-D-aa-tRNA into the ribosomal peptidyl-tRNA binding (P) site. During the next round of continuous translation, however, we find that ribosomes carrying a P-site peptidyl-D-aatRNA partition into subpopulations that are either translationally arrested or that can continue translating. Consistent with its ability to arrest translation, chemical protection experiments and molecular dynamics simulations show that P site-bound peptidyl-D-aa-tRNA can trap the ribosomal peptidyl-transferase center in a conformation in which peptidyl transfer is impaired. Our results reveal a novel mechanism through which D-aa-tRNAs interfere with translation, provide insight into how the TM might be engineered to use D-aa-tRNAs, and increase our understanding of the physiological role of a widely distributed enzyme that clears D-aa-tRNAs from cells.

Original languageEnglish (US)
Pages (from-to)6038-6043
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume112
Issue number19
DOIs
StatePublished - May 12 2015

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Keywords

  • D-amino acids
  • D-aminoacyl-tRNA deacylase
  • Ribosome
  • Translation arrest

ASJC Scopus subject areas

  • General

Cite this

Englander, M. T., Avins, J. L., Fleisher, R. C., Liu, B., Effraim, P. R., Wang, J., Schulten, K., Leyh, T. S., Gonzalez, R. L., & Cornish, V. W. (2015). The ribosome can discriminate the chirality of amino acids within its peptidyl-transferase center. Proceedings of the National Academy of Sciences of the United States of America, 112(19), 6038-6043. https://doi.org/10.1073/pnas.1424712112