Molecular cloning, structural analysis and modelling of the AcAFP antifungal peptide from Aspergillus clavatus.
An abundantly secreted thermostable peptide (designed AcAFP) with a molecular mass of 5777 Da was isolated and purified in a previous work from a local strain of A. clavatus (VR1). Based on the N-terminal amino acid (aa) sequence of the AcAFP peptide, an oligonucleotide probe was derived and allowed the amplification of the encoding cDNA by RT-PCR. This cDNA fragment encodes a pre-pro-protein of 94 aa which appears to be processed to a mature product of 51 aa cys-rich protein. The deduced aa sequence of the pre-pro-sequence reveals high similarity with ascomycetes antifungal peptide. Comparison of the nucleotide sequence of the genomic fragment and the cDNA clone revealed the presence of an open reading frame of 282 bp interrupted by two small introns of 89 and 56 bp with conserved splice site. The three-dimensional (3D) structure modeling of AcAFP exhibits a compact structure consisting of five anti-parallel beta barrel stabilized by four internal disulfide bridges. The folding pattern revealed also a cationic site and spatially adjacent hydrophobic stretch. The antifungal mechanism was investigated by transmission and confocal microscopy. AcAFP cause cell wall altering in a dose-dependent manner against the phytopathogenic fungus Fusarium oxysporum.
Skouri-Gargouri
Houda
H
Laboratoire de Génétique Moléculaire des Eucaryotes, Centre de Biotechnologie de Sfax, Route Sidi Mansour, BP K 3038-Sfax, Tunisia.
Ben Ali
Mamdouh
M
Gargouri
Ali
A
D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000737:chemistry* / Q000494:pharmacology; D001230:Aspergillus / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D005670:Fusarium / Q000187:drug effects / Q000648:ultrastructure; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000235:genetics* / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid
D000935:Antifungal Agents; D010455:Peptides