FTIR analysis of the interaction of azide with horse heart myoglobin variants.
Abstract: The interaction of azide with variants of horse heart myoglobin (Mb) has been characterized by Fourier transform infrared (FTIR), electron paramagnetic resonance (EPR), and UV-VIS absorption spectroscopy and by molecular modeling calculations. Distal histidine variants (His64Thr, His64Ile, His64Lys) and charged surface variants (Val67Arg, Lys45Glu, Lys45Glu/Lys63Glu) were included in this study. All variants, with the exception of Val67Arg, have a lower azide affinity than the wild-type protein. Analysis of the temperature dependence of the FTIR spectra (277-313 K) revealed that the wild-type protein and all variants exhibit a high-spin/low-spin equilibrium. Introduction of positively charged amino acid residues shifts nu max for the low-spin form to higher energy while negatively charged residues shifted this maximum to lower energy. The low azide binding affinity exhibited by the His64Thr and His64Ile variants is accompanied by a shift of the nu max for the low-spin infrared band to lower energy and by a significant increase in the corresponding half-bandwidths. This observation indicates greater mobility of the bound azide ligand in these variants. The His64Lys variant exhibits two infrared bands attributable to low-spin forms that are assigned to two different conformations of the lysyl residue. In one conformation, the lysine is proposed to form a hydrogen bond with the bound azide similar to that proposed to occur between the distal histidine and bound azide, and in the other conformation no interaction occurs.
Publication Date: 1994-06-21 PubMed ID: 8011626DOI: 10.1021/bi00190a013Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The research investigates the effect of different variants of horse heart myoglobin (a type of protein) on their interaction with azide, using techniques such as Fourier transform infrared (FTIR) spectroscopy and molecular modelling calculations. The study finds that most variants of the protein, except one, exhibit lower azide affinity compared to the wild-type protein, and the nature of amino acid residues influences the energy levels associated with the low-spin form of the protein.
Understanding the Research
- The central aim of this research is to understand the interaction of azide, a negatively charged ion, with various versions of horse heart myoglobin, a protein responsible for transporting oxygen in muscle cells.
- Different scientific techniques were applied, including Fourier Transform Infrared (FTIR) spectroscopy, Electron Paramagnetic Resonance (EPR), Ultraviolet-Visible (UV-VIS) absorption spectroscopy, and molecular modelling computations.
- These techniques are essential in characterising the behaviour and structural changes in proteins when they interact with other chemical entities such as azide.
The Variants Considered in the Study
- The researchers explored numerous myoglobin variants (distal histidine variants and charged surface variants).
- All these variants, except for Val67Arg, demonstrated a lower affinity for azide compared to the wild-type protein. Affinity refers to the ability of the protein to bind with azide.
The Findings of the Research
- Analysing the temperature-dependence of the FTIR spectra (277-313 K), the researchers learned that the wild-type protein and all the variants presented a high-spin/low-spin equilibrium.
- The introduction of positively charged amino acid residues pushed the maximum energy (nu max) for the low-spin form to higher energy, while negatively charged residues moved the maximum to lower energy.
- For the variants His64Thr and His64Ile, low azide binding affinity was observed, along with a shift of the nu max for the low-spin infrared band to lower energy. This indicates that there is more mobility in the bound azide ligand in these variants compared to the others.
- The His64Lys variant showcased two infrared bands attributed to low-spin forms, indicating two differing conformations of the lysyl residue. This shows that this variant has a unique behaviour when interacting with azide, depending on how the lysine residue is arranged.
Implications and Significance of the Research
- Understanding these interactions is vital for further developing our comprehension of how proteins work and how modifications to their structure can affect their properties and functions.
- The findings could have applications in the design of protein-based biosensors or biocatalysts where interaction with certain ions (like azide) is necessary.
- It could also lead to more targeted biopharmaceutical research where understanding the impacts of amino acid changes in protein structure can provide insights into treating protein-related diseases.
Cite This Article
APA
Bogumil R, Hunter CL, Maurus R, Tang HL, Lee H, Lloyd E, Brayer GD, Smith M, Mauk AG.
(1994).
FTIR analysis of the interaction of azide with horse heart myoglobin variants.
Biochemistry, 33(24), 7600-7608.
https://doi.org/10.1021/bi00190a013 Publication
Researcher Affiliations
- Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, Canada.
MeSH Terms
- Animals
- Azides / metabolism
- Binding Sites
- Electrochemistry
- Electron Spin Resonance Spectroscopy
- Heme / metabolism
- Histidine / chemistry
- Horses
- Models, Molecular
- Molecular Structure
- Mutation
- Myocardium / chemistry
- Myoglobin / chemistry
- Myoglobin / metabolism
- Protein Binding
- Recombinant Proteins / chemistry
- Recombinant Proteins / metabolism
- Spectrophotometry
- Spectroscopy, Fourier Transform Infrared
- Structure-Activity Relationship
- Temperature
Citations
This article has been cited 4 times.- Cooper IB, Barry BA. Azide as a probe of proton transfer reactions in photosynthetic oxygen evolution.. Biophys J 2008 Dec 15;95(12):5843-50.
- Zahran ZN, Chooback L, Copeland DM, West AH, Richter-Addo GB. Crystal structures of manganese- and cobalt-substituted myoglobin in complex with NO and nitrite reveal unusual ligand conformations.. J Inorg Biochem 2008 Feb;102(2):216-33.
- Hoy JA, Smagghe BJ, Halder P, Hargrove MS. Covalent heme attachment in Synechocystis hemoglobin is required to prevent ferrous heme dissociation.. Protein Sci 2007 Feb;16(2):250-60.
- Maurus R, Bogumil R, Nguyen NT, Mauk AG, Brayer G. Structural and spectroscopic studies of azide complexes of horse heart myoglobin and the His-64-->Thr variant.. Biochem J 1998 May 15;332 ( Pt 1)(Pt 1):67-74.
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