The sequence-specific assignment of the 1H-NMR spectrum of an enzyme, horse-muscle acylphosphatase.
Abstract: A complete range of two-dimensional NMR experiments was used for the assignment of the 1H-NMR spectrum of horse muscle acylphosphatase. Firstly the spin systems of some easily identifiable amino acid side chains were assigned. These side chains involved all the aromatic residues and all the leucine, valine, isoleucine, threonine, alanine, proline as well as some of the glycine residues. Analysis of nuclear Overhauser enhancement spectra in our previous work had identified the sequential and long-range patterns characteristics for secondary structure elements. This result had also provided the identification of the main-chain alpha and amide proton resonances. Several of the completely assigned spin systems were then identified as being part of the secondary structure units which led, after analysis of the primary amino acid sequence, to unambiguous sequence-specific assignments. The identification and assignment of the remaining side-chain resonances was then completed and are reported here. These results provide a complete data base for the three-dimensional structure determination of this enzyme in solution.
Publication Date: 1989-06-01 PubMed ID: 2543576DOI: 10.1111/j.1432-1033.1989.tb14803.xGoogle Scholar: Lookup
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- Journal Article
Summary
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This research focuses on the use of two-dimensional nuclear magnetic resonance (NMR) to understand the structure of an enzyme, horse-muscle acylphosphatase. It provides a detailed evaluation of specific amino acid chains and lays the groundwork for a complete three-dimensional structure of the enzyme.
Method of Research
- The researchers used a complete range of two-dimensional NMR experiments, a technique that uses the magnetic properties of atomic nuclei to identify the structure of molecules.
- They focused their analysis on several particular types of amino acid side chains that are easily identifiable, including all the aromatic residues and all the leucine, valine, isoleucine, threonine, alanine, proline, and some of the glycine residues.
Previous Findings
- In past research, the scientists established patterns from nuclear Overhauser enhancement spectra; this is a technique in NMR spectroscopy that reveals spatial and temporal correlations between spins in molecules.
- This effort enabled them to identify the features of specific secondary structure elements.
- The result of this previous work had also helped in the identification of the alpha and amide proton resonances in the main-chain.
New Findings
- In this study, some of the fully assigned spin systems, which are sets of protons that interact with each other, were found to be part of secondary structural units. This discovery aligns with an analysis of the primary amino acid sequence.
- This led to unambiguous sequence-specific assignments, which are crucial for determining the structure of the enzyme.
- The team then completed the identification and assignment of the remaining side-chain resonances.
Conclusion
- The results of this study have provided a comprehensive database for three-dimensional structural determination of the horse muscle acylphosphatase enzyme in its natural, liquid state.
- This is significant for understanding the functioning of the enzyme, with potential implications for biochemical and biomedical research.
Cite This Article
APA
Saudek V, Boyd J, Williams RJ, Stefani M, Ramponi G.
(1989).
The sequence-specific assignment of the 1H-NMR spectrum of an enzyme, horse-muscle acylphosphatase.
Eur J Biochem, 182(1), 85-93.
https://doi.org/10.1111/j.1432-1033.1989.tb14803.x Publication
Researcher Affiliations
- Inorganic Chemistry Laboratory, University of Oxford, England.
MeSH Terms
- Acid Anhydride Hydrolases
- Amino Acid Sequence
- Amino Acids / analysis
- Animals
- Deuterium
- Horses
- Hydrogen Bonding
- Magnetic Resonance Spectroscopy / methods
- Muscles / enzymology
- Phosphoric Monoester Hydrolases / analysis
- Protein Conformation
Citations
This article has been cited 7 times.- Chiti F, Taddei N, Stefani M, Dobson CM, Ramponi G. Reduction of the amyloidogenicity of a protein by specific binding of ligands to the native conformation. Protein Sci 2001 Apr;10(4):879-86.
- Chiti F, Taddei N, Bucciantini M, White P, Ramponi G, Dobson CM. Mutational analysis of the propensity for amyloid formation by a globular protein. EMBO J 2000 Apr 3;19(7):1441-9.
- Modesti A, Taddei N, Chiti F, Bucciantini M, Magherini F, Rigacci S, Stefani M, Raugei G, Ramponi G. Properties of Cys21-mutated muscle acylphosphatases. J Protein Chem 1996 Jan;15(1):27-34.
- Pazzagli L, Cappugi G, Camici G, Manao G, Ramponi G. Bovine testis acylphosphatase: purification and amino acid sequence. J Protein Chem 1993 Oct;12(5):593-601.
- Paoli P, Camici G, Manao G, Ramponi G. 2-Methoxybenzoyl phosphate: a new substrate for continuous fluorimetric and spectrophotometric acyl phosphatase assays. Experientia 1995 Jan 15;51(1):57-62.
- Stefani M, Degl'Innocenti D, Berti A, Cappugi G, Manao G, Camici G, Ramponi G. Purification and characterization of acylphosphatase erythrocyte isoenzyme from turkey muscle. J Protein Chem 1990 Oct;9(5):633-40.
- Berti A, Tremori E, Pazzagli L, Degl'Innocenti D, Camici G, Cappugi G, Manao G, Ramponi G. Rat muscle acylphosphatase: purification, amino sequence, and immunological characterization. J Protein Chem 1991 Feb;10(1):91-102.
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