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Azidohomoalanine: A Minimally Invasive, Versatile, and Sensitive Infrared Label in Proteins To Study Ligand Binding


Zanobini, Claudio; Bozovic, Olga; Jankovic, Brankica; Koziol, Klemens L; Johnson, Philip J M; Hamm, Peter; Gulzar, Adnan; Wolf, Steffen; Stock, Gerhard (2018). Azidohomoalanine: A Minimally Invasive, Versatile, and Sensitive Infrared Label in Proteins To Study Ligand Binding. Journal of Physical Chemistry B, 122(44):10118-10125.

Abstract

The noncanonical amino acid azidohomoalanine (Aha) is known to be an environment-sensitive infrared probe for the site-specific investigation of protein structure and dynamics. Here, the capability of that label is explored to detect protein ligand interactions by incorporating it in the vicinity of the binding groove of a PDZ2 domain. Circular dichroism and isothermal titration calorimetry measurements reveal that the perturbation of the protein system by mutation is negligible, with minimal influence on protein stability and binding affinity. Two-dimensional infrared spectra exhibit small (1-3 cm(-1)) but clearly measurable red shifts of the Aha vibrational frequency upon binding of two different peptide ligands, while accompanying molecular dynamics simulations suggest that these red shifts are induced by polar contacts with side chains of the peptide ligands. Hence, Aha is a versatile and minimally invasive vibrational label that is not only able to report on large structural changes during, e.g., protein folding, but also on very subtle changes of the electrostatic environment upon ligand binding.

Abstract

The noncanonical amino acid azidohomoalanine (Aha) is known to be an environment-sensitive infrared probe for the site-specific investigation of protein structure and dynamics. Here, the capability of that label is explored to detect protein ligand interactions by incorporating it in the vicinity of the binding groove of a PDZ2 domain. Circular dichroism and isothermal titration calorimetry measurements reveal that the perturbation of the protein system by mutation is negligible, with minimal influence on protein stability and binding affinity. Two-dimensional infrared spectra exhibit small (1-3 cm(-1)) but clearly measurable red shifts of the Aha vibrational frequency upon binding of two different peptide ligands, while accompanying molecular dynamics simulations suggest that these red shifts are induced by polar contacts with side chains of the peptide ligands. Hence, Aha is a versatile and minimally invasive vibrational label that is not only able to report on large structural changes during, e.g., protein folding, but also on very subtle changes of the electrostatic environment upon ligand binding.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Department of Chemistry
Dewey Decimal Classification:540 Chemistry
Uncontrolled Keywords:Physical and Theoretical Chemistry, Materials Chemistry, Surfaces, Coatings and Films
Language:English
Date:8 November 2018
Deposited On:05 Feb 2019 12:52
Last Modified:05 Feb 2019 13:06
Publisher:American Chemical Society (ACS)
ISSN:1520-5207
OA Status:Closed
Publisher DOI:https://doi.org/10.1021/acs.jpcb.8b08368
Project Information:
  • : FunderSwiss National Science Foundation (SNF) through the NCCR MUST
  • : Grant ID200021_165789/1
  • : Project Title
  • : FunderDeutsche Forschungsgemeinschaft
  • : Grant IDSTO 247/10-1
  • : Project Title

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