Inhibitory activity against plant pathogenic fungi of extracts from Myoporum bontioides A. Gray and indentification of active ingredients.

BACKGROUND: In order to understand the bioactivity of Myoporum bontioides A. Gray against plant pathogens and determine its active ingredients, the inhibitory activities of methanol extracts from M. bontioides against Fusarium oxysporum f. sp. niveum (E. F. Smith) Snyder & Hansen, Pestalotia mangiferae P. Henn., Thielaviopsis paradoxa (De Seynes) v. Hohnel, Colletotrichum musae (Berk. & M. A. Curtis) v. Arx, Alternaria alternata (Fr.) Keissler, Mycosphaerella sentina (Fr.) Schroter and Sphaceloma fawcettii Jenk. were evaluated using a growth rate method, and the active ingredient was isolated by activity-directed isolation and identified by determination and analysis of IR, (1)H NMR, (13)C NMR and mass spectra and correlative physical constants. RESULTS: The results showed that the extracts from stems and leaves of M. bontioides exhibited inhibitory activity against the seven fungi, with > 58% inhibition at 10 g L(-1) after 72 h. The active compound was isolated and identified as (-)-epingaione, and showed inhibitory activity against the above seven fungi. The inhibitory activity against P. mangiferae was the highest, with an EC(50) value of 77 mg L(-1). The EC(50) values against the other six fungi were 147-245 mg L(-1). (-)-Epingaione also inhibited spore germination of F. oxysporum f. sp. niveum, T. paradoxa and S. fawcettii. CONCLUSION: (-)-Epingaione demonstrated broad-spectrum inhibitory activity against plant pathogenic fungi and is promising for exploitation as a fungicide.

Expression, purification, and characterization of human acetyl-CoA carboxylase 2.

The full-length human acetyl-CoA carboxylase 1 (ACC1) was expressed and purified to homogeneity by two separate groups (Y.G. Gu, M. Weitzberg, R.F. Clark, X. Xu, Q. Li, T. Zhang, T.M. Hansen, G. Liu, Z. Xin, X. Wang, T. McNally, H. Camp, B.A. Beutel, H.I. Sham, Synthesis and structure-activity relationships of N-{3-[2-(4-alkoxyphenoxy)thiazol-5-yl]-1-methylprop-2-ynyl}carboxy derivatives as selective acetyl-CoA carboxylase 2 inhibitors, J. Med. Chem. 49 (2006) 3770-3773; D. Cheng, C.H. Chu, L. Chen, J.N. Feder, G.A. Mintier, Y. Wu, J.W. Cook, M.R. Harpel, G.A. Locke, Y. An, J.K. Tamura, Expression, purification, and characterization of human and rat acetyl coenzyme A carboxylase (ACC) isozymes, Protein Expr. Purif., in press). However, neither group was successful in expressing the full-length ACC2 due to issues of solubility and expression levels. The two versions of recombinant human ACC2 in these reports are either truncated (lacking 1-148 aa) or have the N-terminal 275 aa replaced with the corresponding ACC1 region (1-133 aa). Despite the fact that ACC activity was observed in both cases, these constructs are not ideal because the N-terminal region of ACC2 could be important for the correct folding of the catalytic domains. Here, we report the high level expression and purification of full-length human ACC2 that lacks only the N-terminal membrane attachment sequence (1-20 and 1-26 aa, respectively) in Trichoplusia ni cells. In addition, we developed a sensitive HPLC assay to analyze the kinetic parameters of the recombinant enzyme. The recombinant enzyme is a soluble protein and has a K(m) value of 2 microM for acetyl-CoA, almost 30-fold lower than that reported for the truncated human ACC2. Our recombinant enzyme also has a lower K(m) value for ATP (K(m)=52 microM). Although this difference could be ascribed to different assay conditions, our data suggest that the longer human ACC2 produced in our system may have higher affinities for the substrates and could be more similar to the native enzyme.