Structural investigation of pig metmyoglobin by 129Xe NMR spectroscopy.
Abstract: The potentiality of xenon's sensitivity to its local magnetic environment is thoroughly investigated to probe internal structural differences between pig and horse metmyoglobin (MMb). These MMb's differ by 14 amino acids. One of these, Ile142 in horse MMb, is located in the proximal cavity, which is the xenon-binding site in horse MMb, and is replaced by Met142 in pig MMb. Specific and non-specific xenon-protein interactions are investigated here by 129Xe NMR chemical shifts and relaxation rate in aqueous solutions of pig MMb as a function of the xenon and protein concentrations. The results are complemented with 129Xe NMR data of the isostructural carbonmonoxy myoglobin (COMb), with computational calculations in order to highlight the structural differences between the cavities, and 1H NMR spectra to test the dependence of the 1H chemical shift on the addition of xenon. The 129Xe chemical shift NMR parameters are analysed quantitatively in terms of a two-site model. Xenon forms a 1:1 complex with the protein, characterized by an equilibrium binding constant K=[Xe]in/([Xe]out[MMb]), and exchanges rapidly between a cavity within the protein (X(ein)) and all other environments (Xe(out)). A comparison of equilibrium constant, K (74 M(-1)) in pig and K (146 M(-1)) in horse, reveals differences in affinity of xenon to the interior of pig MMb. Changes in xenon binding in both pig and horse MMb are also pointed out by other experimental results, e.g. the difference in the estimated delta(in), which is shifted downfield in pig MMb and upfield in horse MMb, with respect to 129Xe in buffer solution; the xenon-iron distance, 7.4 A, which is longer in the pig than was found in the horse, 5.3 A.
Publication Date: 2004-09-18 PubMed ID: 15374622DOI: 10.1016/j.bbagen.2004.06.011Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research investigates the potential of xenon’s sensitivity to its local magnetic environment to understand structural differences between pig and horse metmyoglobin. The study explored xenon protein interactions using multiple techniques and discovered discrepancies in xenon’s affinity to the interior of pig metmyoglobin in comparison with that of the horse.
Context and Purpose of the Research
- The research aimed to utilize the magnetic sensitivity of xenon to probe the internal structural differences between pig and horse metmyoglobin (MMb).
- Xenon was chosen due to both its specific and non-specific interactions with proteins.
- Distinguishing these internal differences could provide valuable insights into the behaviour and characteristics of proteins.
Methodology
- The research methods involved the application of 129Xe NMR chemical shifts and relaxation rate in aqueous solutions of pig MMb as a function of both the xenon and protein concentrations.
- The results were then augmented with 129Xe NMR data of the isostructural carbonmonoxy myoglobin (COMb) and computational calculations to outline the structural differences between the cavities.
- A two-site model was utilized for a quantitative analysis of the 129Xe chemical shift NRM parameters.
- Additional tests were conducted using 1H NMR spectra to evaluate the reliance of the 1H chemical shift on the introduction of xenon.
Key Findings
- Xenon was found to form a 1:1 complex with the protein, where it quickly exchanges between a cavity within the protein and all other environments.
- There was a discrepancy in the equilibrium constant in the pig (74 M(-1)) and horse (146 M(-1)), which suggested differences in the affinity of xenon with the interior of pig MMb.
- A change in xenon binding in both pig and horse MMb was confirmed by other experimental results. For instance, the difference in the estimated delta(in), which shifted downwards in pig MMb and upwards in horse MMb, compared to 129Xe in buffer solution.
- The distance between xenon and iron was lengthier in pig (7.4 A) than in horse (5.3 A).
Significance of the Research
- The research offers a novel approach to analyze the structure of proteins using the magnetic properties of xenon.
- The findings highlight the noteworthy differences in the affinity of xenon to the interior of pig and horse metmyoglobin, thereby enhancing our understanding of the structure and behavior of proteins.
Cite This Article
APA
Corda M, Era B, Fais A, Casu M.
(2004).
Structural investigation of pig metmyoglobin by 129Xe NMR spectroscopy.
Biochim Biophys Acta, 1674(2), 182-192.
https://doi.org/10.1016/j.bbagen.2004.06.011 Publication
Researcher Affiliations
- Dipartimento di Scienze Applicate ai Biosistemi, Cittadella Universitaria di Monserrato, 09042 Monserrato, CA, Italy.
MeSH Terms
- Animals
- Binding Sites
- Horses
- Magnetics
- Metmyoglobin / chemistry
- Metmyoglobin / genetics
- Metmyoglobin / metabolism
- Myoglobin / chemistry
- Nuclear Magnetic Resonance, Biomolecular
- Protein Binding
- Protein Conformation
- Swine
- Xenon Isotopes / chemistry
- Xenon Isotopes / metabolism
Citations
This article has been cited 2 times.- Roose BW, Zemerov SD, Wang Y, Kasimova MA, Carnevale V, Dmochowski IJ. A Structural Basis for (129) Xe Hyper-CEST Signal in TEM-1 β-Lactamase. Chemphyschem 2019 Jan 21;20(2):260-267.
- Roose BW, Zemerov SD, Dmochowski IJ. Xenon-Protein Interactions: Characterization by X-Ray Crystallography and Hyper-CEST NMR. Methods Enzymol 2018;602:249-272.
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