Factors affecting the yield and properties of bacterial cellulose

Acetobacter xylinum E₂₅ has been applied in our studies in order to find optimal culture conditions for effective bacterial cellulose (BC) production. The strain displays significantly higher stability in BC production under stationary culture conditions. In contrast, intensive agitation and aeration appear to drastically reduce cellulose synthesis since such conditions induced formation of spontaneous cellulose nonproducing mutants (Cel−), which dominated in the culture. Mutation frequency strictly depends on the medium composition in agitated cultures. Enrichment of the standard SH and Yamanaka media with 1% ethanol significantly enhanced BC production in stationary cultures. Horizontal fermentors equipped with rotating discs or rollers were successfully applied in order to improve culture conditions. Relatively slow rotation velocity (4 rpm) and large surface area enabling effective cell attachment are optimal parameters for cellulose production. Physical properties of BC samples synthesized either in stationary cultures or in a horizontal fermentor revealed that cellulose from stationary cultures demonstrated a much higher value of Young's modulus, but a much lower value of water-holding capacity. Journal of Industrial Microbiology & Biotechnology (2002) 29, 189–195 doi:10.1038/sj.jim.7000303 Received 01 March 2002/ Accepted in revised form 18 July 2002     

Interaction of hepatitis C virus proteins with host cell membranes and lipids

For replication, viruses depend on specific components and energy supplies from the host cell. The main steps in the lifecycle of positive-strand RNA viruses depend on cellular membranes. Interest is increasing in studying the interactions between host cell membranes and viral proteins to understand how such viruses replicate their genome and produce infectious particles. These studies should also lead to a better knowledge of the different mechanisms underlying membrane-protein associations. The various molecular interactions of hepatitis C virus proteins with the membranes and lipids of the infected cell highlight how a virus can exploit the diversity of interactions that occur between proteins and membranes or lipid structures.     

Synthesis, and functional properties of a modified human insulin A-chain: implication in a 'mini-insulin' structure determination

The design and total synthesis of a novel insulin A-chain mutant, analogue 3, is reported. In this compound, the cysteines implied in the two insulin inter-chain disulfide bridges are replaced by two serines (residues Ser(A7) and Ser(A20)) and the intra-A-chain disulfide bridge (residues Cys(A6) and Cys(A11)) is conserved. This A-chain analogue (3) has been tested in three in vitro cell culture assays, using insulin as a reference. The data clearly showed that analogue 3 mimics insulin effects on DNA synthesis, glucose uptake and glycogen synthesis without loss of potency as compared to insulin. To our knowledge, these are the first results showing that an isolated insulin chain displays functional properties similar to those of insulin. The implication of these new findings in insulin structure-function relationships and in a 'mini-insulin' structure determination is discussed.     

A newborn infant with sternal malformation/vascular dysplasia association

Sternal malformation/vascular dysplasia complex was described by Hersch et al. in 1985. The principle findings include cleft of the sternum covered by an atrophic skin, a midline abdominal raphe and hemangiomatosis. The inheritance pattern seems to be sporadic. We report a newborn baby with sternal defect, cleft lip and palate, supraumbilical raphe and hemangiomas.     

Effect of surfactant on pulmonary expression of type IIA PLA(2) in an animal model of acute lung injury

We previously showed that the seminatural surfactant Curosurf inhibits the in vitro synthesis of secretory type IIA phospholipase A(2) (sPLA(2)-IIA) in alveolar macrophages (AM). These cells are the main source of sPLA(2)-IIA in a guinea pig model of lipopolysaccharide (LPS)-induced acute lung injury (ALI). Here, we investigate the effect of Curosurf on the pulmonary synthesis of sPLA(2)-IIA in this ALI model. Our results showed that intratracheal administration of LPS (330 microg/kg) induced an increase in pulmonary expression of sPLA(2)-IIA, which was inhibited when animals received Curosurf (16 mg/guinea pig) 30 min or 8 h after LPS instillation. When AM were isolated from LPS-treated animals and cultured in conditioned medium, they expressed higher levels of sPLA(2)-IIA than AM from saline-treated animals. This ex vivo sPLA(2)-IIA expression was significantly reduced when guinea pigs received Curosurf 30 min after LPS instillation. Finally, we examined the effect of Curosurf on pulmonary inflammation measured 8 or 24 h after LPS administration. Curosurf instillation 30 min or 8 h after LPS reversed the increase in tumor necrosis factor-alpha expression, polymorphonuclear cell extravasation, and protein concentration in bronchoalveolar lavage fluids. Curosurf also decreased the bronchial reactivity induced by LPS. We conclude that Curosurf inhibits the pulmonary expression of sPLA(2)-IIA and exhibits palliative anti-inflammatory effects in an animal model of ALI.     

IRT1, an Arabidopsis transporter essential for iron uptake from the soil and for plant growth

Plants are the principal source of iron in most diets, yet iron availability often limits plant growth. In response to iron deficiency, Arabidopsis roots induce the expression of the divalent cation transporter IRT1. Here, we present genetic evidence that IRT1 is essential for the uptake of iron from the soil. An Arabidopsis knockout mutant in IRT1 is chlorotic and has a severe growth defect in soil, leading to death. This defect is rescued by the exogenous application of iron. The mutant plants do not take up iron and fail to accumulate other divalent cations in low-iron conditions. IRT1-green fluorescent protein fusion, transiently expressed in culture cells, localized to the plasma membrane. We also show, through promoter::beta-glucuronidase analysis and in situ hybridization, that IRT1 is expressed in the external cell layers of the root, specifically in response to iron starvation. These results clearly demonstrate that IRT1 is the major transporter responsible for high-affinity metal uptake under iron deficiency.     

Biogenesis of nanotubular network in Toxoplasma parasitophorous vacuole induced by parasite proteins

The intracellular parasite Toxoplasma gondii develops within a nonfusogenic vacuole containing a network of elongated nanotubules that form connections with the vacuolar membrane. Parasite secretory proteins discharged from dense granules (known as GRA proteins) decorate this intravacuolar network after invasion. Herein, we show using specific gene knockout mutants, that the unique nanotubule conformation of the network is induced by the parasite secretory protein GRA2 and further stabilized by GRA6. The vacuolar compartment generated by GRA2 knockout parasites was dramatically disorganized, and the normally tubular network was replaced by small aggregated material. The defect observed in Deltagra2 parasites was evident from the initial stages of network formation when a prominent cluster of multilamellar vesicles forms at a posterior invagination of the parasite. The secretory protein GRA6 failed to localize properly to this posterior organizing center in Deltagra2 cells, indicating that this early conformation is essential to proper assembly of the network. Construction of a Deltagra6 mutant also led to an altered mature network characterized by small vesicles instead of elongated nanotubules; however, the initial formation of the posterior organizing center was normal. Complementation of the Deltagra2 knockout with mutated forms of GRA2 showed that the integrity of both amphipathic alpha-helices of the protein is required for correct formation of the network. The induction of nanotubues by the parasite protein GRA2 may be a conserved feature of amphipathic alpha-helical regions, which have also been implicated in the organization of Golgi nanotubules and endocytic vesicles in mammalian cells.