Structural flexibility of the macrophage dengue virus receptor CLEC5A: implications for ligand binding and signaling

The human C-type lectin-like molecule CLEC5A is a critical macrophage receptor for dengue virus. The binding of dengue virus to CLEC5A triggers signaling through the associated adapter molecule DAP12, stimulating proinflammatory cytokine release. We have crystallized an informative ensemble of CLEC5A structural conformers at 1.9-Å resolution and demonstrate how an on-off extension to a β-sheet acts as a binary switch regulating the flexibility of the molecule. This structural information together with molecular dynamics simulations suggests a mechanism whereby extracellular events may be transmitted through the membrane and influence DAP12 signaling. We demonstrate that CLEC5A is homodimeric at the cell surface and binds to dengue virus serotypes 1–4. We used blotting experiments, surface analyses, glycan microarray, and docking studies to investigate the ligand binding potential of CLEC5A with particular respect to dengue virus. This study provides a rational foundation for understanding the dengue virus-macrophage interaction and the role of CLEC5A in dengue virus-induced lethal disease.     

Molecular Evidence of <Emphasis Type="Italic">Wolbachia</Emphasis> Infection in Natural Populations of Tropical Odonates

Wolbachia are endosymbiotic bacteria that cause reproductive alterations in numerous arthropod species. Using a PCR-based method, we found that, out of 33 odonate species, four species were infected with Wolbachia. This finding represents the first record of Wolbachia infection in tropical odonates. Identical wsp gene sequences were found in the Wolbachia-infected common odonate species, Agriocnemis f. femina, collected from different locations in Thailand. The infection frequencies in several natural populations suggest that replacement of uninfected populations by Wolbachia-infected ones has recently occurred in this damselfly species.Received: 12 November 2002 / Accepted: 13 December 2002     

Molecular phylogeny of tephritid fruit flies in the Bactrocera tau complex using the mitochondrial COI sequences.

We compared sequences of the mitochondrial cytochrome oxidase I gene of eight species of the Bactrocera tau complex using Bactrocera dorsalis, Bactrocera pyrifoliae, Ceratitis capitata, Anopheles gambiae, and Locusta migratoria as outgroups. A 639-bp variable region was sequenced. The sequence divergence between species in the B. tau complex ranged from 0.06 to 28%, and up to 29% between the complex and its tephritid outgroups, B. dorsalis and C. capitata. According to the phylogenetic relationships, these members of the B. tau complex could be classified into four clades. Thus, species A and D form clades 1 and 3, respectively, while species C and I belong to clade 4. However, species B, E, and F form a distinct group, clade 2, and infested the fruits of non-cucurbit hosts. Host-plant shifts resulting in adaptive radiation and premating isolation among species might play an important role in species differentiation of the B. tau complex.     

Fed-batch optimization of alpha-amylase and protease-producing Bacillus subtilis using Markov chain methods

A stoichiometry-based model for the fed-batch culture of the recombinant bacterium Bacillus subtilis ATCC 6051a, producing extracellular alpha-amylase as a desirable product and proteases as undesirable products, was developed and verified. The model was then used for optimizing the feeding schedule in fed-batch culture. To handle higher-order model equations (14 state variables), an optimization methodology for the dual-enzyme system is proposed by integrating Pontryagin's optimum principle with fermentation measurements. Markov chain Monte Carlo (MCMC) procedures were appropriate for model parameter and decision variable estimation by using a priori parameter distributions reflecting the experimental results. Using a simplified Metropolis-Hastings algorithm, the specific productivity of alpha-amylase was maximized and the optimum path was confirmed by experimentation. The optimization process predicted a further 14% improvement of alpha-amylase productivity that could not be realized because of the onset of sporulation. Among the decision variables, the switching time from batch to fed-batch operation (t(s)) was the most sensitive decision variable.