A 2S albumin-homologous protein from passion fruit seeds inhibits the fungal growth and acidification of the medium by Fusarium oxysporum.
Antimicrobial proteins have been isolated from a wide range of plant species. More recently, it has become increasingly clear that these types of proteins play an important role in the protection of plants. In this study, we investigate the presence of defense-related proteins from passion fruit (Passiflora edulis f. flavicarpa) seeds. Initially, seed flour was extracted for 2h (at 4 degrees C) with phosphate buffer, pH 5.5. The precipitate obtained between 0 and 70% relative ammonium sulfate saturation was re-dissolved in distilled water and heated at 80 degrees C for 15 min. The resulting suspension was clarified by centrifugation and the supernatant (F/0-70) was extensively dialyzed. A Sephadex G-50 size exclusion column was employed for further separation of proteins. The fraction with antifungal activity was pooled and submitted to CM-Sepharose cation exchange. Two proteins, named Pf1 and Pf2, were eluted in 0.1 and 0.2M of salt, respectively, and submitted to reverse-phase chromatography in HPLC. This fraction inhibited the growth, in an in vitro assay, of the phytopathogenic fungi Fusarium oxysporum and colletotrichum lindemuthianum and the yeast Saccharomyces cerevisiae and strongly inhibited glucose-stimulated acidification of the medium by F. oxysporum in a dose-dependent manner. The molecular masses of these proteins, referred to now as Pf1-RP and Pf2-RP, were obtained by MALDI-TOF spectrometry and corresponded to 12,088 Da for Pf1-RP and 11,930 Da for Pf2-RP. These proteins were also subjected to automated N-terminal amino acid sequencing. Sequence comparisons for the heavy subunit of Pf2-RP showed the presence of a protein with a high degree of homology to storage 2S albumins.
Agizzio
Ana Paula
AP
Laboratório de Fisiologia e Bioquímica de Microrganismos Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense, Campos dos Goytacazes, RJ 28013-600, Brazil.
Carvalho
André O
AO
Ribeiro
Suzanna de Fátima F
Sde F
Machado
Olga L T
OL
Alves
Elias W
EW
Okorokov
Lev A
LA
Samarão
Solange S
SS
Bloch
Carlos
C
Prates
Maura V
MV
Gomes
Valdirene M
VM
D055516:2S Albumins, Plant; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D003470:Culture Media / Q000737:chemistry; D005670:Fusarium / Q000737:chemistry / Q000187:drug effects* / Q000254:growth & development* / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D029598:Passiflora / Q000737:chemistry / Q000378:metabolism* / Q000382:microbiology; D010940:Plant Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D012639:Seeds / Q000737:chemistry / Q000378:metabolism / Q000382:microbiology; D016415:Sequence Alignment; D013045:Species Specificity
D055516:2S Albumins, Plant; D052179:Antigens, Plant; D003470:Culture Media; D010940:Plant Proteins; D005947:Glucose