Determination of the disulfide array of the first inducible antifungal peptide from insects: drosomycin from Drosophila melanogaster.
Drosomycin is a 44-residue antifungal peptide with four intramolecular disulfide bridges which have been isolated from immune-challenged Drosophila. To produce adequate amounts of this peptide for 3D-structure analysis, studies on the mode of action and activity spectrum, we expressed a synthetic cDNA in Saccharomyces cerevisiae. For this purpose, we used the mating factor alpha gene and concomitantly overexpressed the KEX2 gene to increase the yield of fully processed drosomycin. Using a combination of Edman degradation and mass spectrometry, we show that drosomycin shares the same array of intramolecular disulfide bridges than plant defensins, in addition to their sequence similarities.
Michaut
L
L
Institut de Biologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique, Réponse Immunitaire et Développement chezles Insectes, Strasbourg, France.
Fehlbaum
P
P
Moniatte
M
M
Van Dorsselaer
A
A
Reichhart
J M
JM
Bulet
P
P
D000595:Amino Acid Sequence; D000818:Animals; D000935:Antifungal Agents / Q000737:chemistry* / Q000302:isolation & purification; D001483:Base Sequence; D004274:DNA, Recombinant; D004220:Disulfides / Q000737:chemistry*; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000737:chemistry*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D019476:Insect Proteins; D000072235:Mating Factor; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000235:genetics; D043484:Proprotein Convertases; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D013381:Subtilisins / Q000235:genetics
D000935:Antifungal Agents; D004274:DNA, Recombinant; D004220:Disulfides; D029721:Drosophila Proteins; D019476:Insect Proteins; D010455:Peptides; D011506:Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C091227:DRS protein, Drosophila; D000072235:Mating Factor; D043484:Proprotein Convertases; D013381:Subtilisins; C058751:KEX2 protein, S cerevisiae