Trehalose-6-phosphate synthase from the cat flea Ctenocephalides felis and Drosophila melanogaster: gene identification, cloning, heterologous functional expression and identification of inhibitors by high throughput screening

被引:29
作者
Kern, C. [1 ]
Wolf, C. [1 ]
Bender, F. [1 ]
Berger, M. [1 ]
Noack, S. [1 ]
Schmalz, S. [1 ]
Ilg, T. [1 ]
机构
[1] Intervet Innovat GmbH, D-55270 Zur Propstei, Schwabenheim, Germany
关键词
trehalose; insecticide target; pesticide target; high throughput screening; ectoparasite; TREHALOSE-PHOSPHATE-SYNTHASE; ESCHERICHIA-COLI; BIOSYNTHESIS; METABOLISM; TREHAZOLIN; PROTEINS; ANHYDROBIOSIS; ECTOPARASITES; ACCUMULATION; STRATEGIES;
D O I
10.1111/j.1365-2583.2012.01151.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Trehalose phosphate synthase (EC 2.4.1.15; TPS) is the crucial enzyme for the biosynthesis of trehalose, the main haemolymph sugar of insects, and therefore a potential insecticidal molecular target. In this study, we report the functional heterologous expression of Drosophila melanogaster TPS, the gene identification, full length cDNA cloning and functional expression of cat flea (Ctenocephalides felis) TPS, and the MichaelisMenten constants for their specific substrates glucose-6-phosphate and uridinediphosphate-glucose. A novel high throughput screening-compatible TPS assay and its use for the identification of the first potent insect TPS inhibitors from a large synthetic compound collection (>115 000 compounds) is described. One compound class that emerged in this screening, the 4-substituted 2,6-diamino-3,5-dicyano-4H-thiopyrans, was further investigated by analysing preliminary structureactivity relationships. Here, compounds were identified that show low mu M to high nM half maximal inhibitory concentrations on insect TPS and that may serve as lead compounds for the development of insecticides with a novel mode of action.
引用
收藏
页码:456 / 471
页数:16
相关论文
共 73 条
[2]   Enhanced thermotolerance for ethanol fermentation of Saccharomyces cerevisiae strain by overexpression of the gene coding for trehalose-6-phosphate synthase [J].
An, Ming-Zhe ;
Tang, Yue-Qin ;
Mitsumasu, Kanako ;
Liu, Ze-Shen ;
Shigeru, Morimura ;
Kenji, Kida .
BIOTECHNOLOGY LETTERS, 2011, 33 (07) :1367-1374
[3]   TREHAZOLIN, A SLOW, TIGHT-BINDING INHIBITOR OF SILKWORM TREHALASE [J].
ANDO, O ;
NAKAJIMA, M ;
KIFUNE, M ;
FANG, H ;
TANZAWA, K .
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS, 1995, 1244 (2-3) :295-302
[4]   TREHAZOLIN, A NEW TREHALASE INHIBITOR [J].
ANDO, O ;
SATAKE, H ;
ITOI, K ;
SATO, A ;
NAKAJIMA, M ;
TAKAHASHI, S ;
HARUYAMA, H ;
OHKUMA, Y ;
KINOSHITA, T ;
ENOKITA, R .
JOURNAL OF ANTIBIOTICS, 1991, 44 (10) :1165-1168
[5]  
[Anonymous], 2012, Molecular Cloning: A Laboratory Manual
[6]   Physiological roles of trehalose in bacteria and yeasts:: a comparative analysis [J].
Argüelles, JC .
ARCHIVES OF MICROBIOLOGY, 2000, 174 (04) :217-224
[7]   TREHALASE INHIBITORS, VALIDOXYLAMINE-A AND RELATED-COMPOUNDS AS INSECTICIDES [J].
ASANO, N ;
TAKEUCHI, M ;
KAMEDA, Y ;
MATSUI, K ;
KONO, Y .
JOURNAL OF ANTIBIOTICS, 1990, 43 (06) :722-726
[8]   The regulation of trehalose metabolism in insects [J].
Becker, A ;
Schloder, P ;
Steele, JE ;
Wegener, G .
EXPERIENTIA, 1996, 52 (05) :433-439
[9]   The role of trehalose in the physiology of nematodes [J].
Behm, CA .
INTERNATIONAL JOURNAL FOR PARASITOLOGY, 1997, 27 (02) :215-229
[10]   Trehalose accumulation during cellular stress protects cells and cellular proteins from damage by oxygen radicals [J].
Benaroudj, N ;
Lee, DH ;
Goldberg, AL .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2001, 276 (26) :24261-24267