Dataset Information

Structural and functional analyses of a sterol carrier protein in Spodoptera litura.

ABSTRACT:

Backgrounds

In insects, cholesterol is one of the membrane components in cells and a precursor of ecdysteroid biosynthesis. Because insects lack two key enzymes, squalene synthase and lanosterol synthase, in the cholesterol biosynthesis pathway, they cannot autonomously synthesize cholesterol de novo from simple compounds and therefore have to obtain sterols from their diet. Sterol carrier protein (SCP) is a cholesterol-binding protein responsible for cholesterol absorption and transport.

Results

In this study, a model of the three-dimensional structure of SlSCPx-2 in Spodoptera litura, a destructive polyphagous agricultural pest insect in tropical and subtropical areas, was constructed. Docking of sterol and fatty acid ligands to SlSCPx-2 and ANS fluorescent replacement assay showed that SlSCPx-2 was able to bind with relatively high affinities to cholesterol, stearic acid, linoleic acid, stigmasterol, oleic acid, palmitic acid and arachidonate, implying that SlSCPx may play an important role in absorption and transport of these cholesterol and fatty acids from host plants. Site-directed mutation assay of SlSCPx-2 suggests that amino acid residues F53, W66, F89, F110, I115, T128 and Q131 are critical for the ligand-binding activity of the SlSCPx-2 protein. Virtual ligand screening resulted in identification of several lead compounds which are potential inhibitors of SlSCPx-2. Bioassay for inhibitory effect of five selected compounds showed that AH-487/41731687, AG-664/14117324, AG-205/36813059 and AG-205/07775053 inhibited the growth of S. litura larvae.

Conclusions

Compounds AH-487/41731687, AG-664/14117324, AG-205/36813059 and AG-205/07775053 selected based on structural modeling showed binding affinity to SlSCPx-2 protein and inhibitory effect on the growth of S. litura larvae.

SUBMITTER: Zhang L

PROVIDER: S-EPMC3893073 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Publications

Structural and functional analyses of a sterol carrier protein in Spodoptera litura.

Zhang Lili L Li Ding D Xu Rui R Zheng Sichun S He Hongwu H Wan Jian J Feng Qili Q

PloS one 20140115 1

<h4>Backgrounds</h4>In insects, cholesterol is one of the membrane components in cells and a precursor of ecdysteroid biosynthesis. Because insects lack two key enzymes, squalene synthase and lanosterol synthase, in the cholesterol biosynthesis pathway, they cannot autonomously synthesize cholesterol de novo from simple compounds and therefore have to obtain sterols from their diet. Sterol carrier protein (SCP) is a cholesterol-binding protein responsible for cholesterol absorption and transport ...[more]

PMID: 24454688

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Project description:BackgroundSterol carrier protein-2/3-oxoacyl-CoA thiolase (SCPx) gene has been suggested to be involved in absorption and transport of cholesterol. Cholesterol is a membrane component and is a precursor of ecdysteroids, but cannot be synthesized de novo in insects. However, a direct association between SCPx gene expression, cholesterol absorption and development in lepidopteran insects remains to be experimentally demonstrated.ResultsAn SCPx cDNA (SlSCPx) cloned from the common cutworm, Spodoptera litura, was characterized. The SlSCPx cDNA encoded a 535-amino acid protein consisting of a 3-oxoacyl-CoA thiolase (SCPx-t) domain and a SCP-2 (SCPx-2) domain. SlSCPx mRNA was expressed predominately in the midgut, while SlSCPx-2 mRNA was detected in the midgut, fat body and epidermis and no SlSCPx-t mRNA was detected. A 58-kDa full-length SCPx protein and a 44-kDa SCPx-t protein were detected in the midgut of sixth instar larvae when the anti-SlSCPx-t antibody was used in western blotting analysis; a 16-kDa SCP-2 protein was detected when anti-SlSCPx-2 antibody was used. SlSCPx protein was post-translationally cleaved into two smaller proteins, SCPx-t and SCPx-2. The gene appeared to be expressed into two forms of mRNA transcripts, which were translated into the two proteins, respectively. SlSCPx-t and SlSCPx-2 proteins have distinct and different locations in the midgut of sixth instar larvae. SlSCPx and SlSCPx-t proteins were detected predominately in the cytoplasm, whereas SlSCPx-2 protein was detected in the cytoplasm and nuclei in the Spli-221 cells. Over-expression of SlSCPx and SlSCPx-2 proteins enhanced cholesterol uptake into the Spli-221 cells. Knocking-down SlSCPx transcripts by dsRNA interference resulted in a decrease in cholesterol level in the hemolymph and delayed the larval to pupal transition.ConclusionSpatial and temporal expression pattern of this SlSCPx gene during the larval developmental stages of S. litura showed its specific association with the midgut at the feeding stage. Over-expression of this gene increased cholesterol uptake and interference of its transcript decreased cholesterol uptake and delayed the larval to pupal metamorphosis. All of these results taken together suggest that this midgut-specific SlSCPx gene is important for cholesterol uptake and normal development in S. litura.

Dataset Information

Structural and functional analyses of a sterol carrier protein in Spodoptera litura.

Backgrounds

Results

Conclusions

Publications

Structural and functional analyses of a sterol carrier protein in Spodoptera litura.

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