Molten globule structure of equine beta-lactoglobulin probed by hydrogen exchange.
Abstract: The molten globule structure of equine beta-lactoglobulin has been inferred from the hydrogen exchange protection of the backbone amide protons. In order to make it possible to measure the hydrogen exchange kinetics of the individual backbone amide protons, the uniformly (15)N-labeled recombinant protein was expressed in Escherichia coli and the NMR peak assignment was obtained for most of the backbone protons. The chemical shift and NOE results obtained under the condition where the protein assumes the native structure are fully consistent with the known secondary structure of bovine beta-lactoglobulin, indicating that the equine protein has a similar native conformation to that of the bovine protein. The hydrogen exchange rate of the individual backbone amide protons was measured under the conditions where the protein assumes the native and molten globule states. In the native state, strong protection was observed for the residues located in the eight (A to H) strands, which form a barrel structure, and residues of a major helix. In the molten globule state at acidic pH conditions, significant protection from the exchange has been observed for residues located in the A, F, G and H strands in the native structure. The pattern of protection is consistent with a native-like beta-sheet formation by these strands. The residues located in a major helix of the native structure are also protected, suggesting that this helix is formed in the molten globule and is packed against the sheet as in the native structure. These results indicate that a native-like subdomain is formed in the molten globule state of equine beta-lactoglobulin.
Copyright 2000 Academic Press.
Publication Date: 2000-06-03 PubMed ID: 10835282DOI: 10.1006/jmbi.2000.3761Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The study analysed the structure of a specific protein in horseshoes, called equine beta-lactoglobulin, using a technique called hydrogen exchange. With the help of this technique, researchers were able to identify certain protective measures of the protein structure, in both its native and molten states, providing insights into its composition and functions.
Methodology
- The researchers began by expressing a uniformly (15)N-labeled recombinant protein in E. coli and obtained nuclear magnetic resonance (NMR) peak assignments for most of the backbone protons. This was done to measure the hydrogen exchange kinetics of these protons.
- They tested the protein under conditions that allowed it to assume both its native structure and a less-orderly state known as the molten globule state.
- They then observed the rates of hydrogen exchange under these varying conditions.
Findings
- The researchers found that when the protein was in its native conformation, there was a strong protection for the residues located in the eight strands (labelled A to H) which form a barrel structure, and a major helix.
- Under the conditions where the protein was in a molten globule state, a significant protection was observed for residues located in the A, F, G, and H strands of the native structure. This protection indicated a native-like beta-sheet formation by these strands.
- In the molten globule state, residues from a major helix of the native structure were also protected, suggesting that this helix is formed in the molten globule and is packed against the sheet as in the native structure.
Conclusion
- The study concluded that a native-like subdomain is formed in the molten globule state of equine beta-lactoglobulin. This suggests that even when the protein is in a less-ordered state, it retains some structural characteristics of its native conformation.
- This research gives insights into protein structure and function in different states, which can be valuable for understanding protein-related biomedical processes and developing related treatments.
Cite This Article
APA
Kobayashi T, Ikeguchi M, Sugai S.
(2000).
Molten globule structure of equine beta-lactoglobulin probed by hydrogen exchange.
J Mol Biol, 299(3), 757-770.
https://doi.org/10.1006/jmbi.2000.3761 Publication
Researcher Affiliations
- Department of Bioengineering Faculty of Engineering, Soka University, Tokyo, Hachioji, 192-8577, Japan.
MeSH Terms
- Amides / metabolism
- Amino Acid Sequence
- Animals
- Cattle
- Circular Dichroism
- Deuterium / metabolism
- Horses
- Hydrogen / metabolism
- Hydrogen-Ion Concentration
- Kinetics
- Lactoglobulins / chemistry
- Lactoglobulins / genetics
- Lactoglobulins / metabolism
- Models, Molecular
- Molecular Sequence Data
- Nuclear Magnetic Resonance, Biomolecular
- Protein Folding
- Protein Structure, Secondary
- Protons
- Recombinant Proteins / chemistry
- Recombinant Proteins / genetics
- Recombinant Proteins / metabolism
Citations
This article has been cited 9 times.- Sawyer L. β-Lactoglobulin and Glycodelin: Two Sides of the Same Coin?. Front Physiol 2021;12:678080.
- Baliga C, Selmke B, Worobiew I, Borbat P, Sarma SP, Trommer WE, Varadarajan R, Aghera N. CcdB at pH 4 Forms a Partially Unfolded State with a Dry Core.. Biophys J 2019 Mar 5;116(5):807-817.
- Ikeguchi M. Transient non-native helix formation during the folding of β-lactoglobulin.. Biomolecules 2014 Feb 13;4(1):202-16.
- Matsumura Y, Shinjo M, Matsui T, Ichimura K, Song J, Kihara H. Structural study of hNck2 SH3 domain protein in solution by circular dichroism and X-ray solution scattering.. Biophys Chem 2013 May-Jun;175-176:39-46.
- Ohtomo H, Konuma T, Utsunoiya H, Tsuge H, Ikeguchi M. Structure and stability of Gyuba, a β-lactoglobulin chimera.. Protein Sci 2011 Nov;20(11):1867-75.
- Gasymov OK, Abduragimov AR, Glasgow BJ. Molten globule state of tear lipocalin: ANS binding restores tertiary interactions.. Biochem Biophys Res Commun 2007 Jun 1;357(2):499-504.
- Alcaraz LA, Jiménez B, Moratal JM, Donaire A. An NMR view of the unfolding process of rusticyanin: Structural elements that maintain the architecture of a beta-barrel metalloprotein.. Protein Sci 2005 Jul;14(7):1710-22.
- Fujiwara K, Ikeguchi M, Sugai S. A partially unfolded state of equine beta-lactoglobulin at pH 8.7.. J Protein Chem 2001 Feb;20(2):131-7.
- Carrotta R, Bauer R, Waninge R, Rischel C. Conformational characterization of oligomeric intermediates and aggregates in beta-lactoglobulin heat aggregation.. Protein Sci 2001 Jul;10(7):1312-8.
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