Complete amino acid analysis of peptides and proteins after hydrolysis by a mixture of sepharose-bound peptidases

Incubation with a mixture of Sepharose-bound peptidases was shown to result in the quantitative release of amino acids from certain peptides and S-aminoethylated proteins. Subtraction of the low background values of amino acids generated by the enzymes enables amino acid ratios of corticotrophin-(1-...

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Bibliographic Details
Published in:Biochemical journal 1972-09, Vol.129 (3), p.695-701
Main Authors: Bennett, H P, Elliott, D F, Evans, B E, Lowry, P J, McMartin, C
Format: Article
Language:English
Subjects:
Adrenocorticotropic Hormone - analysis
Amino Acids - analysis
Aminopeptidases
Angiotensin II - analysis
Bradykinin - analysis
Chymotrypsin
Dipeptidases
Hydrolysis
Muramidase - analysis
Peptide Hydrolases
Peptides - analysis
Polysaccharides
Protein Binding
Proteins - analysis
Ribonucleases - analysis
Trypsin
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Summary:Incubation with a mixture of Sepharose-bound peptidases was shown to result in the quantitative release of amino acids from certain peptides and S-aminoethylated proteins. Subtraction of the low background values of amino acids generated by the enzymes enables amino acid ratios of corticotrophin-(1-24)-tetracosapeptide to be determined with a standard deviation on repeat digestions of 3-5%. Good values were obtained for amino acids that are completely or partially destroyed on acid hydrolysis, i.e. tryptophan, tyrosine, serine, asparagine and glutamine. Experiments with peptides containing d-amino acids showed that the enzyme mixture is stereospecific and could therefore be used to detect the presence of d-residues in peptides. The enzyme mixture completely hydrolyses peptide fragments obtained after Edman degradation and should therefore be useful for determining sequences of peptides containing acid-labile amino acid residues. The activities of the bound enzymes were unaltered over a period of 7 months and they provide a simple, reproducible procedure for the quantitative determination of amino acids in peptides and proteins containing l-amino acids.
ISSN:0264-6021
0306-3283
1470-8728
DOI:10.1042/bj1290695