Determination of iodide using flow injection with acidic potassium permanganate chemiluminescence detection.

A simple and rapid flow-injection method is described for the determination of iodide, based on potassium permanganate chemiluminescence detection via oxidation of formaldehyde in aqueous hydrochloric acid. The calibration graph was linear over the range 1.0-12 x 10(-6) mol/L (r2 = 0.9955) with relative standard deviations (n = 4) in the range 1.0-3.5%. The detection limit (3sigma) was 1.0 x 10(-7) mol/L, with sample throughput of 120/h. The effect of interfering cations [Ca(II), Mg(II), Ni(II), Fe(II), Fe(III) and Pb(II)] and anions (Cl-, SO4(2-), PO4(3-), NO3-, NO2-, F- and SO3(2-)) were studied. The method was applied to iodized salt samples and the results obtained in the range 0.03 +/- 0.005 - 0.10 +/- 0.006 mg I/g were in reasonable agreement with the amount labelled. The method was statistically compared with the results obtained by titration; no significant disagreement at 95% confidence was observed.     

Bioaugmentation and coexistence of two functionally similar bacterial strains in aerobic granules

The survival of the inoculated microbial culture is critical for successful bioaugmentation but impossible to predict precisely. As an alternative strategy, bioaugmentation of a group of microorganisms may improve reliability of bioaugmentation. This study evaluated simultaneous bioaugmentation of two functionally similar bacterial strains in aerobic granules. The two strains, Pandoraea sp. PG-01 and Rhodococcus erythropolis PG-03, showed high phenol degradation and growth rates in phenol medium, but they were characterized as having a poor aggregation activity and weak bioflocculant-producing and biofilm-forming abilities. In the spatially homogeneous batch conditions, strain PG-01 with higher growth rates outcompeted strain PG-03. However, the two strains could stably coexist in the spatially heterogeneous conditions. Then the two strains were mixed and bioaugmented into activated sludge in two sequencing batch reactors, which were operated with the different settling times of 5 and 30 min, respectively. Aerobic granules were developed only in the reactor with a settling time of 5 min. Fluorescence in situ hybridization and denaturing gradient gel electrophoresis showed that the two strains could coexist in aerobic granules but not in activated sludge. These findings suggested that the compact structure of aerobic granules provided spatial isolation for coexistence of competitively superior and inferior strains with similar functions.     

Oxidative stress and neurological disorders in relation to blood lead levels in children

Oxidative stress plays a pivotal role in the pathogenesis of neurological disorders. Free radical generation appears to be the mode of lead toxicity. We evaluated the effects of blood lead levels on oxidative stress parameters in children suffering from neurological disorders. Thirty children (aged 3-12 years) with neurological disorders (cerebral palsy [n = 12], seizures [n = 11], and encephalopathy [n = 7]) were recruited in the study group. Sixty healthy children (aged 3-12 years) from similar socio-economic environments and not suffering from any chronic disease were taken as the controls. Blood lead levels and oxidant/antioxidant status were determined. Mean blood lead level was significantly higher while delta-aminolevulinic acid dehydratase (delta-ALAD) activity, a biomarker for lead exposure, was significantly lower in the study group as compared to the control group (P < 0.05 for each). Malondialdehyde (MDA) levels, an end-product of lipid peroxidation, were significantly higher while the antioxidant glutathione (GSH) levels were significantly lower in the study group as compared to the control group (P < 0.05 for each). Activities of the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) were significantly higher in the study group than those of the control group (P < 0.05 for each). There were significant negative correlations of blood lead levels with delta-ALAD (r = -0.35; P < 0.05) and GSH (r = -0.31; P < 0.05), and positive correlations with MDA (r = 0.37; P < 0.05), SOD (r = 0.53; P < 0.05), and CAT (r = 0.31; P < 0.05). In turn, delta-ALAD had significant negative correlations with MDA (r = -0.29; P < 0.05), SOD (r = -0.28; P < 0.05) and CAT (r = -0.34; P < 0.05), but positive correlation with GSH (r = 0.32; P < 0.05). Although a causal pathway can not be determined from the present study, our findings indicate lead-induced oxidative stress in blood of children with neurological disorders. Lead-induced oxidative stress as an underlying mechanism for neurological diseases in children warranted further investigation.     

Analysis of secondary structure predictions of dengue virus type 2 NS2B/NS3 against crystal structure to evaluate the predictive power of the in silico methods

Multiple sequence alignment was performed against eight proteases from the Flaviviridae family using ClustalW to illustrate conserved domains. Two sets of prediction approaches were applied and the results compared. Firstly, secondary structure prediction was performed using available structure prediction servers. The second approach made use of the information on the secondary structures extracted from structure prediction servers, threading techniques and DSSP database of some of the templates used in the threading techniques. Consensus on the one-dimensional secondary structure of Den2 protease was obtained from each approach and evaluated against data from the recently crystallised Den2 NS2B/NS3 obtained from the Protein Data Bank (PDB). Results indicated the second approach to show higher accuracy compared to the use of prediction servers only. Thus, it is plausible that this approach is applicable to the initial stage of structural studies of proteins with low amino acid sequence homology against other available proteins in the PDB.     

Model of cellular mechanotransduction via actin stress fibers

Mechanical stresses due to blood flow regulate vascular endothelial cell structure and function and play a key role in arterial physiology and pathology. In particular, the development of atherosclerosis has been shown to correlate with regions of disturbed blood flow where endothelial cells are round and have a randomly organized cytoskeleton. Thus, deciphering the relation between the mechanical environment, cell structure, and cell function is a key step toward understanding the early development of atherosclerosis. Recent experiments have demonstrated very rapid (\(\sim \)100 ms) and long-distance (\(\sim \)10 \(\upmu \)m) cellular mechanotransduction in which prestressed actin stress fibers play a critical role. Here, we develop a model of mechanical signal transmission within a cell by describing strains in a network of prestressed viscoelastic stress fibers following the application of a force to the cell surface. We find force transmission dynamics that are consistent with experimental results. We also show that the extent of stress fiber alignment and the direction of the applied force relative to this alignment are key determinants of the efficiency of mechanical signal transmission. These results are consistent with the link observed experimentally between cytoskeletal organization, mechanical stress, and cellular responsiveness to stress. Based on these results, we suggest that mechanical strain of actin stress fibers under force constitutes a key link in the mechanotransduction chain.     

High yield lipase-catalyzed synthesis of Engkabang fat esters for the cosmetic industry

Engkabang fat esters were produced via alcoholysis reaction between Engkabang fat and oleyl alcohol, catalyzed by Lipozyme RM IM. The reaction was carried out in a 500 ml Stirred tank reactor using heptane and hexane as solvents. Response surface methodology (RSM) based on a four-factor-five-level Central composite design (CCD) was applied to evaluate the effects of synthesis parameters, namely temperature, substrate molar ratio (oleyl alcohol: Engkabang fat), enzyme amount and impeller speed. The optimum yields of 96.2% and 91.4% were obtained for heptane and hexane at the optimum temperature of 53.9 °C, impeller speeds of 309.5 and 309.0 rpm, enzyme amounts of 4.82 and 5.65 g and substrate molar ratios of 2.94 and 3.39:1, respectively. The actual yields obtained compared well with the predicted values of 100.0% and 91.5%, respectively. Meanwhile, the properties of the esters show that they are suitable to be used as ingredient for cosmetic applications.     

CYP2E1 and Parkinson’s disease in a MPTP-induced C57BL/6 mouse model

Background

A proline-to-serine substitution at position-56 (P56S) of vesicle-associated membrane protein-associated protein B (VAPB) causes a form of dominantly inherited motor neuron disease (MND), including typical and atypical amyotrophic lateral sclerosis (ALS) and a mild late-onset spinal muscular atrophy (SMA). VAPB is an integral endoplasmic reticulum (ER) protein and has been implicated in various cellular processes, including ER stress, the unfolded protein response (UPR) and Ca²⁺ homeostasis. However, it is unclear how the P56S mutation leads to neurodegeneration and muscle atrophy in patients. The formation of abnormal VAPB-positive inclusions by mutant VAPB suggests a possible toxic gain of function as an underlying mechanism. Furthermore, the amount of VAPB protein is reported to be reduced in sporadic ALS patients and mutant SOD1G93A mice, leading to the hypothesis that wild type VAPB plays a role in the pathogenesis of ALS without VAPB mutations.

Results

To investigate the pathogenic mechanism in vivo, we generated human wild type (wtVAPB) and mutant VAPB (muVAPB) transgenic mice that expressed the transgenes broadly in the CNS. We observed robust VAPB-positive aggregates in the spinal cord of muVAPB transgenic mice. However, we failed to find an impairment of motor function and motor neuron degeneration. We also did not detect any change in the endogenous VAPB level or evidence for induction of the unfolded protein response (UPR) and coaggregation of VAPA with muVAPB. Furthermore, we crossed these VAPB transgenic mice with mice that express mutant SOD1G93A and develop motor neuron degeneration. Overexpression of neither wtVAPB nor muVAPB modulated the protein aggregation and disease progression in the SOD1G93A mice.

Conclusion

Overexpression of VAPBP56S mutant to approximately two-fold of the endogenous VAPB in mouse spinal cord produced abundant VAPB aggregates but was not sufficient to cause motor dysfunction or motor neuron degeneration. Furthermore, overexpression of either muVAPB or wtVAPB does not modulate the course of ALS in SOD1G93A mice. These results suggest that changes in wild type VAPB do not play a significant role in ALS cases that are not caused by VAPB mutations. Furthermore, these results suggest that muVAPB aggregates are innocuous and do not cause motor neuron degeneration by a gain-of-toxicity, and therefore, a loss of function may be the underlying mechanism.     

Altered acetate metabolism and biomass production in several Escherichia coli mutants lacking rpoS-dependent metabolic pathway genes

The stress responsive sigma factor RpoS regulates the expression of tktB and talAgenes of the non-oxidative pentose phosphate (PP) pathway, and fumCand acnA genes of the TCA cycle at the stationary phase of growth. In the present study, batch cultivations were performed using tktB, talA, fumC or acnA-knockout mutants of Escherichia coli to observe the metabolic changes at different phases of growth compared to the wild type strain. Although the specific growth rates of the mutants were similar to the wild type, acetate yield was nearly half in all mutants except the acnA mutant. Altered acetate yield in the mutants was also accompanied by variations in the biomass yield. While the biomass yield in both the tktB and talA mutants was increased by 13.8%, biomass was 5.5% and 13.8% lower in the fumC and acnA mutants, respectively. Upregulation of global regulators such as rpoS and soxRS, the acs, aceA, aceB genes, and several TCA cycle genes such as fumC, acnA and sucA, is consistent with higher acetate consumption and biomass yield in the tktB and talA mutants. On the other hand, the fumC and acnA mutants, with their impaired TCA cycles, were unable to utilize acetate for biomass production in spite of the higher expression of rpoS and soxRS.     

The role of ADP-ribosylation factor and SAR1 in vesicular trafficking in plants

Ras-like small GTP binding proteins regulate a wide variety of intracellular signalling and vesicular trafficking pathways in eukaryotic cells including plant cells. They share a common structure that operates as a molecular switch by cycling between active GTP-bound and inactive GDP-bound conformational states. The active GTP-bound state is regulated by guanine nucleotide exchange factors (GEF), which promote the exchange of GDP for GTP. The inactive GDP-bound state is promoted by GTPase-activating proteins (GAPs) which accelerate GTP hydrolysis by orders of magnitude. Two types of small GTP-binding proteins, ADP-ribosylation factor (Arf) and secretion-associated and Ras-related (Sar), are major regulators of vesicle biogenesis in intracellular traffic and are founding members of a growing family that also includes Arf-related proteins (Arp) and Arf-like (Arl) proteins. The most widely involved small GTPase in vesicular trafficking is probably Arf1, which not only controls assembly of COPI- and AP1, AP3, and AP4/clathrin-coated vesicles but also recruits other proteins to membranes, including some that may be components of further coats. Recent molecular, structural and biochemical studies have provided a wealth of detail of the interactions between Arf and the proteins that regulate its activity as well as providing clues for the types of effector molecules which are controlled by Arf. Sar1 functions as a molecular switch to control the assembly of protein coats (COPII) that direct vesicle budding from ER. The crystallographic analysis of Sar1 reveals a number of structurally unique features that dictate its function in COPII vesicle formation. In this review, I will summarize the current knowledge of Arf and Sar regulation in vesicular trafficking in mammalian and yeast cells and will highlight recent advances in identifying the elements involved in vesicle formation in plant cells. Additionally, I will briefly discuss the similarities and dissimilarities of vesicle traffic in plant, mammalian and yeast cells.