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Optimised expression and spectral analysis of the target enzyme CYP51 from Penicillium digitatum with possible new DMI fungicides
Author(s) -
Zhang Jianhua,
Zhao Li,
Zhang Jie,
Han Rui,
Li Shuxiang,
Yuan Yongze,
Wan Jian,
Xiao Wenjing,
Liu Deli
Publication year - 2010
Publication title -
pest management science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.296
H-Index - 125
eISSN - 1526-4998
pISSN - 1526-498X
DOI - 10.1002/ps.2021
Subject(s) - penicillium digitatum , fungicide , azole , escherichia coli , biology , chemistry , horticulture , biochemistry , antifungal , microbiology and biotechnology , gene
BACKGROUND: Sterol 14α‐demethylase (CYP51), a key target of azole (DMI) fungicides, can be expressed in both prokaryotes and eukaryotes. Green mould of citrus, caused by Penicillium digitatum (Pers.) Sacc., is a serious post‐harvest disease. To develop specific and more effective fungicides against this disease, the characteristics of the interaction between sterol 14α‐demethylase from P. digitatum (PdCYP51) and possible new fungicides were analysed. The cyp 51 gene of P. digitatum was cloned and expressed under different conditions in Escherichia coli (Mig.) Cast. & Chalm., and the binding spectra of PdCYP51 were explored by the addition of two commercial azoles and four new nitrogen compounds. RESULTS: The yield of soluble protein (PdCYP51) was largest when expressed in Rosetta (DE3) induced by 0.5 m M IPTG for 8 h at 30 °C. Compound B (7‐methoxy‐2 H ‐benzo[ b ][1,4]thiazine‐3‐amine) showed the strongest binding activity of the four new nitrogen compounds, with a K d value of 0.268 µ M . The K d values of the six compounds were significantly correlated with their EC 50 values. CONCLUSION: The spectral analysis and bioassay results could be used to screen the new chemical entities effectively. Compound B, selected by virtual screening from a commercial chemical library, is a candidate for a new DMI fungicide. These results provide a theoretical basis and new ideas for efficient design and development of new antifungal agents. Copyright © 2010 Society of Chemical Industry