Production of stable isotope enriched antimicrobial peptides in Escherichia coli: an application to the production of a 15N-enriched fragment of lactoferrin.
A method is described for the production of recombinant isotopically enriched peptides in E. coli. Peptides are produced in high yield as fusion proteins with ketosteroid isomerase which form insoluble inclusion bodies. This insoluble form allows easy purification, stabilizes the peptide against degradation and prevents bactericidal activity of the peptide. Cyanogen bromide cleavage released peptide which was conjugated with alkylamines to form lipopeptide. An important advantage of this system is that it allows production of peptides that are toxic to bacteria, which we have demonstrated on a dodecapeptide based on residues 21-31 of human bactericidal protein lactoferrin.
Majerle
A
A
Laboratory for Molecular Modeling and NMR spectroscopy, National Institute of Chemistry, Ljubljana, Slovenia.
Kidric
J
J
Jerala
R
R
D003001:Cloning, Molecular / Q000379:methods; D003488:Cyanogen Bromide; D004926:Escherichia coli; D006801:Humans; D007553:Isotope Labeling / Q000379:methods; D007781:Lactoferrin / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016339:Spectrometry, Mass, Fast Atom Bombardment
D009587:Nitrogen Isotopes; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D007781:Lactoferrin; D003488:Cyanogen Bromide