Enzyme-mediated peptide synthesis using acylpeptide hydrolase.
Acylpeptide hydrolase is shown to catalyse the specific addition of a single amino acid to the N-terminus of a peptide. The stabilised Sepharose-coupled form of the enzyme is used to couple a carboxy-methylated N-formyl (or N-acetyl) amino acid to a short pre-existing peptide. The yield is improved by optimal timing of the reaction and the presence of moderate concentrations (5%) of N,N-dimethylformamide. Two tripeptides, Ac-Ala-Ala-Ala and fMet-Leu-Phe (f, formyl) were synthesized by this technique (in yields of 2% and 0.064% respectively). The products were characterised by HPLC, amino acid analysis, mass spectroscopy and protein sequencing. The synthetic fMet-Leu-Phe also had biological activity, in that it stimulated superoxide generation by granulocytes. Acylpeptide hydrolase could therefore be a very useful tool for the synthesis and modification of peptides.
Farries
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TC
MRC Collaborative Centre, London, England.
Auffret
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AD
Aitken
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D000596:Amino Acids / Q000032:analysis; D000626:Aminopeptidases / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D013058:Mass Spectrometry; D010447:Peptide Hydrolases / Q000378:metabolism*; D010455:Peptides / Q000138:chemical synthesis*
D000596:Amino Acids; D010455:Peptides; D010447:Peptide Hydrolases; D000626:Aminopeptidases; C038220:acylaminoacyl-peptidase