The complete amino acid sequence of horse muscle acylphosphatase.
Abstract: The amino acid sequence of horse muscle acylphosphatase is given in the present paper. The carboxymethylated enzyme consists of a single polypeptide chain of 98 amino acid residues with an acetyl group blocking the NH2 terminus and a tyrosine at the COOH terminus. The calculated molecular weight of the native protein, a mixed disulfide with glutathione, is 11,365. The carboxymethylated protein was cleaved by cyanogen bromide. The three expected fragments were purified; moreover, an additional fragment, derived from a partial failure of cleavage at methionine-24, was purified and characterized. The structures of the cyanogen bromide fragments were established by subfragmentation with endopeptidases, and the sequences of the overlapping subfragments were determined. From the results, it was possible to order the peptides within the sequence and then to establish the complete primary structure of the enzyme.
Publication Date: 1980-07-25 PubMed ID: 6248536
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- Journal Article
Summary
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The research paper presents the detailed amino acid sequence of horse muscle acylphosphatase, a crucial protein enzyme.
Overview of the Study
- The study is primarily concerned with the determination of the complete amino acid sequence of horse muscle acylphosphatase, a critical protein enzyme known to play a role in energy metabolism.
- The researchers found that the carboxymethylated form of this enzyme comprises a single polypeptide chain of 98 amino acids, with an acetyl group blocking one end (the NH2 terminus) and a tyrosine residue at the other end (the COOH terminus).
- The calculated molecular weight of the unmodified protein (considered a mixed disulfide with glutathione) is approximately 11,365.
Methodology
- The researchers used cyanogen bromide to cleave (split) the carboxymethylated protein.
- Post cleavage, the three expected fragments were purified. Simultaneously, an additional fragment, product of a partial cleavage failure at methionine-24, was also purified and characterized.
- Subfragmentation of the cyanogen bromide fragments was carried out using endopeptidases to determine their structure.
- The sequences of these overlapping subfragments were then resolved to identify their correct order.
Findings
- The researchers were able to establish the complete primary structure of the horse muscle acylphosphatase enzyme.
- This was achieved by employing the results from the fragmentation, purification, and sequencing processes, enabling the researchers to correctly order the peptides within the sequence. This is significant because it divulges foundational information about the enzyme structure, which could be beneficial in further studies exploring its functions and interactions within horse muscle metabolism.
Cite This Article
APA
Cappugi G, Manao G, Camici G, Ramponi G.
(1980).
The complete amino acid sequence of horse muscle acylphosphatase.
J Biol Chem, 255(14), 6868-6874.
Publication
Researcher Affiliations
MeSH Terms
- Acid Anhydride Hydrolases
- Amino Acid Sequence
- Aminopeptidases
- Animals
- Carboxypeptidases
- Disulfides / analysis
- Horses
- Molecular Weight
- Muscles / enzymology
- Organophosphates
- Peptide Fragments / analysis
- Phosphoric Monoester Hydrolases
Citations
This article has been cited 8 times.- Paoli P, Fiaschi T, Cirri P, Camici G, Manao G, Cappugi G, Raugei G, Moneti G, Ramponi G. Mechanism of acylphosphatase inactivation by Woodward's reagent K. Biochem J 1997 Dec 15;328 ( Pt 3)(Pt 3):855-61.
- 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.
- Caselli A, Pazzagli L, Paoli P, Manao G, Camici G, Cappugi G, Ramponi G. Porcine liver low M(r) phosphotyrosine protein phosphatase: the amino acid sequence. J Protein Chem 1994 Jan;13(1):107-15.
- 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.
- Liguri G, Nassi P, Camici G, Manao G, Cappugi G, Stefani M, Berti A, Ramponi G. Studies on synthesis and degradation rates and some molecular properties of guinea-pig muscle acylphosphatase. Biochem J 1984 Jan 15;217(2):499-505.
- Manao G, Cappugi G, Modesti A, Stefani M, Marzocchini R, Degl'Innocenti D, Camici G. Guinea pig acylphosphatase: the amino acid sequence. J Protein Chem 1988 Aug;7(4):417-26.
- 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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