Recombinant expression and range of activity of penaeidins, antimicrobial peptides from penaeid shrimp.
Penaeidins are 5.5- to 6.6-kDa antimicrobial peptides recently isolated from the plasma and haemocytes of the tropical shrimp Penaeus vannamei. These molecules differ from the other classes of antimicrobial peptides in that they are composed of a proline-rich N-terminus and of a C-terminus containing six cysteine residues engaged in three disulfide bridges. In order to gain information on their antimicrobial activity, two penaeidins (Pen-2 and Pen-3a) were expressed in Saccharomyces cerevisiae. The recombinant Pen-2 and -3a were characterized in terms of primary structure by Edman degradation, mass spectrometry and gas chromatography. A protocol was then established to purify the amount of penaeidins required for the determination of their activity spectrum. We demonstrate in this study that expression in yeast is appropriate for the large-scale production of functional penaeidins, whose activities are almost indistinguishable from those of the native molecules. Data on Pen-2 and -3a activity demonstrate that penaeidins have a broad spectrum of antifungal properties associated with a fungicidal activity, and that their antibacterial activities are essentially directed against Gram-positive bacteria, with a strain-specific inhibition mechanism. Despite a better efficiency of Pen-3a on most of the tested strains, similar activity spectra and inhibition mechanisms were observed for both Pen-2 and -3a. Finally, no synergistic effect could be observed between the two molecules.
Destoumieux
D
D
IFREMER/CNRS/Université de Montpellier 2, UMR 219, France.
Bulet
P
P
Strub
J M
JM
Van Dorsselaer
A
A
Bachère
E
E
D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D001483:Base Sequence; D002849:Chromatography, Gas; D002851:Chromatography, High Pressure Liquid; D003445:Crustacea; D003545:Cysteine / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010455:Peptides; D010957:Plasmids / Q000378:metabolism; D011392:Proline / Q000737:chemistry; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors
D000900:Anti-Bacterial Agents; D018076:DNA, Complementary; D010455:Peptides; D011506:Proteins; D011994:Recombinant Proteins; C109157:penaeidin 2; C109158:penaeidin 3; D011392:Proline; D003545:Cysteine