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Crystal structures of apo wild-type M. jannaschii tyrosyl-tRNA synthetase (TyrRS) and an engineered TyrRS specific for O-methyl-L-tyrosine

Academic Article
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Overview

related to degree

  • Zhang, Yan Jessie, Ph.D. in Biology, Scripps Research 2001 - 2004

authors

  • Zhang, Yan Jessie
  • Wang, L.
  • Schultz, Peter
  • Wilson, Ian

publication date

  • May 2005

journal

  • Protein Science  Journal

abstract

  • The Methanococcus jannaschii tRNA(Tyr)/TyrRS pair has been engineered to incorporate unnatural amino acids into proteins in E. coli. To reveal the structural basis for the altered specificity of mutant TyrRS for O-methyl-L-tyrosine (OMeTyr), the crystal structures for the apo wild-type and mutant M. jannaschii TyrRS were determined at 2.66 and 3.0 A, respectively, for comparison with the published structure of TyrRS complexed with tRNA(Tyr) and substrate tyrosine. A large conformational change was found for the anticodon recognition loop 257-263 of wild-type TyrRS upon tRNA binding in order to facilitate recognition of G34 of the anticodon loop through pi-stacking and hydrogen bonding interactions. Loop 133-143, which is close to the tRNA acceptor stem-binding site, also appears to be stabilized by interaction with the tRNA(Tyr). Binding of the substrate tyrosine results in subtle and cooperative movements of the side chains within the tyrosine-binding pocket. In the OMeTyr-specific mutant synthetase structure, the signature motif KMSKS loop and acceptor stem-binding loop 133-143 were surprisingly ordered in the absence of bound ATP and tRNA. The active-site mutations result in altered hydrogen bonding and steric interactions which favor binding of OMeTyr over L-tyrosine. The structure of the mutant and wild-type TyrRS now provide a basis for generating new active-site libraries to evolve synthetases specific for other unnatural amino acids.

subject areas

  • Crystallography, X-Ray
  • Methanococcus
  • Methyltyrosines
  • Models, Molecular
  • Protein Conformation
  • Protein Engineering
  • Substrate Specificity
  • Tyrosine-tRNA Ligase
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Research

keywords

  • crystallography
  • protein engineering
  • tyrosyl-tRNA synthetase
  • unnatural amino acid
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Identity

PubMed Central ID

  • PMC2253270

International Standard Serial Number (ISSN)

  • 0961-8368

Digital Object Identifier (DOI)

  • 10.1110/ps.041239305

PubMed ID

  • 15840835
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Additional Document Info

start page

  • 1340

end page

  • 1349

volume

  • 14

issue

  • 5

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