Cloning and expression of pathogenesis-related protein 4 from jelly fig (Ficus awkeotsang Makino) achenes associated with ribonuclease, chitinase and anti-fungal activities

• Published in: Plant physiology and biochemistry Vol. 56; pp. 1 - 13
• Main Authors: Lu, Hsi-Chi, Lin, Jia-Hui, Chua, Anna C.N., Chung, Tse-Yu, Tsai, I.-Chun, Tzen, Jason T.C., Chou, Wing-Ming
• Format: Journal Article
• Language: English
• Published: Paris Elsevier Masson SAS 01.07.2012; Elsevier
• Subjects: Achene; Amino Acid Sequence; Anti-fungal; Antifungal Agents - metabolism; Antifungal Agents - pharmacology; Biological and medical sciences; Chitin - metabolism; Chitinase; Chitinases - metabolism; Cloning, Molecular; DNA, Complementary; Ficus; Ficus - chemistry; Ficus awkeotsang; Fundamental and applied biological sciences. Psychology; Fungi - drug effects; Fusarium - drug effects; Fusarium oxysporum; Gene Expression; Heat treatments; Hot Temperature; Molecular Sequence Data; Pathogenesis-related protein 4; Pathogenesis-related proteins; Pericarp; Pichia - metabolism; Pichia pastoris; Plant physiology and development; Plant Proteins - genetics; Plant Proteins - metabolism; Plant Proteins - pharmacology; Polymerase chain reaction; Recombinants; Ribonuclease; Ribonucleases - genetics; Ribonucleases - metabolism; Ribonucleases - pharmacology; RNA - metabolism; Sclerotium rolfsii; Seeds - chemistry; Thermal stability; ChtBD; Pathogenesis-related protein 4; Anti-fungal; Achene; MALDI/MS; Pericarp; Ficus awkeotsang; Ribonuclease; PR-4; Chitinase; Enzyme; Esterases; Moraceae; Woody plant; Dry fruit; Pathogenesis related protein; Biological activity; Glycosylases; Antifungal agent; Plant physiology; Ficus; Dicotyledones; Glycosidases; Angiospermae; Hydrolases; Spermatophyta
• ISSN: 0981-9428; 1873-2690
• DOI: 10.1016/j.plaphy.2012.04.004

A cDNA fragment (FaPR4) encoding a class I pathogenesis-related protein 4 (PR-4) from Ficus awkeotsang was obtained by PCR cloning. Plant PR-4s were grouped into class I and II, differing by the presence of ChtBD and hinge. The predicted mature FaPR4 comprises N-terminal chitin-binding domain (ChtBD), hinge, Barwin domain and C-terminal extension. FaPR4-C, an N-terminal truncated form of FaPR4, was designed to mimic the structural feature of class II PR-4s. FaPR4 and FaPR4-C were over-expressed in yeast Pichia pastoris, and both recombinants exhibited RNase and anti-fungal activities. To our knowledge, it is the first report that FaPR4, a member of class I PR-4s has RNase activity as class II. FaPR4 possesses better anti-fungal activities toward Fusarium oxysporum and Sclerotium rolfsii than FaPR4-C. Heat-treated FaPR4 remained RNase and anti-fungal activities; while heat-treated FaPR4-C lost those activities. Therefore, ChtBD of FaPR4 may not only contribute to its anti-fungal but also improve the thermal stability of protein. It also implied the correlation of RNase activity with anti-fungal activity of FaPR4-C. Furthermore, FaPR4 was detected to have weak but significant chitinase activity, and its chitinase activity was reduced after heat treatment. The chitinase activity by FaPR4-C was much lower than FaPR4. ► FaPR4 coding for a class I PR-4 from jelly fig achenes was obtained by PCR. ► FaPR4 and FaPR4-C represent class I and II PR-4, each were expressed in yeast. ► FaPR4, heat-treated FaPR4 and FaPR4-C showed RNAase activity. ► FaPR4, heat-treated FaPR4 and FaPR4-C exhibited anti-fungal activity. ► First report indicates that a class I PR-4 has RNase activity.