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871.
Human V(H) domains are promising molecules in applications involving antibodies, in particular, immunotherapy because of their human origin. However, they are, in general, prone to aggregation. Therefore, various strategies have been employed to acquire monomeric human V(H)s. We had previously discovered that filamentous phages displaying engineered monomeric V(H) domains gave rise to significantly larger plaques on bacterial lawns than phages displaying wild type V(H)s with aggregation tendencies. Using plaque size as the selection criterion and a phage-displayed na?ve human V(H) library we identified 15 V(H)s that were monomeric. Additionally, the V(H)s demonstrated good expression yields, good refolding properties following thermal denaturation, resistance to aggregation during long incubation at 37 degrees C, and to trypsin at 37 degrees C. These 15 V(H)s should serve as good scaffolds for developing immunotherapeutics, and the selection method employed here should have general utility for isolating proteins with desirable biophysical properties.  相似文献   
872.
873.
Peroxiredoxins are a large family of peroxidases that have important antioxidant and cell signaling functions. Genes encoding two novel 2-cysteine peroxiredoxin proteins were identified in the expressed sequence tag data base of the helminth parasite Schistosoma mansoni, a causative agent of schistosomiasis. The recombinant proteins showed peroxidase activity in vitro with a variety of hydroperoxides and used both the thioredoxin and the glutathione systems as electron donors. Steady-state kinetic analysis indicated that the new peroxiredoxins had saturable kinetics, whereas a previously identified schistosome peroxiredoxin was found to function with more typical unsaturable (ping-pong) kinetics. The catalytic efficiencies S. mansoni peroxiredoxins were similar to those for other peroxiredoxins studied (10(4)-10(5) m(-1) s(-1)). Mutagenesis of S. mansoni peroxiredoxins indicated that glutathione dependence and kinetic differences were conferred by the C-terminal alpha-helix forming 22 amino acids. This is the first report of 2-cysteine peroxiredoxins efficiently utilizing reducing equivalents from both the thioredoxin and glutathione systems. Studies to determine the resistance to oxidative inactivation, important in regulating cell signaling pathways, showed that S. mansoni possess both bacterial-like resistant and mammalian-like sensitive peroxiredoxins. The susceptibility to oxidative inactivation was conferred by the C-terminal tail containing a tyrosine-phenylalanine motif. S. mansoni is the first organism shown to possess both robust and sensitive peroxiredoxins. The ability of schistosome peroxiredoxins to use alternative electron donors, and their variable resistance to overoxidation may reflect their presence in different cellular sites and emphasizes the significant differences in overall redox balance mechanisms between the parasite and its mammalian host.  相似文献   
874.
The temperature-induced, over-expression of the human growth hormone gene in a recombinant E. coli during high cell density cultivation is reported. Human growth hormone (hGH) production and stability were tested under different heat shock conditions. Cell densities were 25 and 60 g l(-1) in a pH-stat fed-batch mode in defined and complex medium, respectively, and the fermentation time was decreased from 41 to 32 h. hGH was produced at 2 g l(-1) in complex medium. By using glycerol as main carbon source in the complex medium with exponential feeding, cell density and hGH production were increased to 100 g l(-1) and 2.7 g l(-1), respectively.  相似文献   
875.
The recent consensus is that virtually all aspects of response selectivity exhibited by the primary visual cortex are either created or sharpened by cortical inhibitory interneurons. Experimental studies have shown that there are cortical inhibitory cells that are driven by geniculate cells and that, like their cortical excitatory counterparts, are orientation selective, though less sharply tuned. The main goal of this article is to demonstrate how orientation-selective inhibition might be created by the circuitry of the primary visual cortex (striate cortex, V1) from its nonoriented geniculate inputs. To fulfill this goal, first, a Bayes–Markov computational model is developed for the V1 area dedicated to foveal vision. The developed model consists of three parts: (i) a two-layered hierarchical Markov random field that is assumed to generate the activity patterns of the geniculate and cortical inhibitory cells, (ii) a Bayesian computational goal that is formulated based on the maximum a posteriori (MAP) estimation principle, and (iii) an iterative, deterministic, parallel algorithm that leads the cortical circuitry to achieve its assigned computational goal. The developed model is not fully LGN driven and it is not implementable by the neural machinery of V1. The model, then, is transformed into a fully LGN-driven and physiologically plausible form. Computer simulation is used to demonstrate the performance of the developed models.  相似文献   
876.
Nucleotide sequences of the intergenic region between ORF 62 and ORF 63 of equine herpesvirus 1 (EHV-1) isolates were analyzed. The sequences of this region consisted of variable and conserved domains among EHV-1 isolates. An EHV-1 mutant, Ab4-GFP, was constructed by inserting a green fluorescent protein (GFP) expression cassette flanked by lox P at both ends into the intergenic region between ORF 62 and ORF 63. Another mutant, Ab4-loxP, which contains one lox P site, was constructed by excision of the GFP cassette from the Ab4-GFP virus genome by cre enzyme. The recombinant Ab4-GFP formed smaller plaques than the wild type in MDBK cells. Virus production also decreased for Ab4-GFP in multistep growth analyses. Virulence of Ab4-GFP in both mice and hamsters was weaker than that of the wild type. Ab4-loxP exhibited properties similar to those of the wild type. These results suggest that the intergenic region between ORF 62 and ORF 63 plays various roles in the virus growth.  相似文献   
877.
The C-terminal region in class Alpha glutathione transferase A1-1 (GSTA1-1), which forms an amphipathic alpha-helix (helix 9), is known to contribute to the catalytic and non-substrate ligand-binding functions of the enzyme. The region in the apo protein is proposed to be disordered which, upon ligand binding at the active-site, becomes structured and localised. Because Ile219 plays a pivotal role in the stability and localisation of the region, the role of tertiary interactions mediated by Ile219 in determining the conformation and dynamics of the C-terminal region were studied. Ligand-binding microcalorimetric and X-ray structural data were obtained to characterise ligand binding at the active-site and the associated localisation of the C-terminal region. In the crystal structure of the I219A hGSTA1-1.S-hexylglutathione complex, the C-terminal region of one chain is mobile and not observed (unresolved electron density), whereas the corresponding region of the other chain is localised and structured as a result of crystal packing interactions. In solution, the mutant C-terminal region of both chains in the complex is mobile and delocalised resulting in a hydrated, less hydrophobic active-site and a reduction in the affinity of the protein for S-hexylglutathione. Complete dehydration of the active-site, important for maintaining the highly reactive thiolate form of glutathione, requires the binding of ligands and the subsequent localisation of the C-terminal region. Thermodynamic data demonstrate that the mobile C-terminal region in apo hGSTA1-1 is structured and does not undergo ligand-induced folding. Its close proximity to the surface of the wild-type protein is indicated by the concurrence between the observed heat capacity change of complex formation and the type and amount of surface area that becomes buried at the ligand-protein interface when the C-terminal region in the apo protein assumes the same localised structure as that observed in the wild-type complex.  相似文献   
878.
879.

Few studies have investigated the association between sleep pattern and nutrient intake pattern. This study was conducted to examine the associations between patterns of nutrient intake and sleep pattern. 108 overweight and obese individuals were recruited to participate in the present cross-sectional study. Participant underwent sleep evaluation through ActiGraph. A 3-day food dietary record was obtained to estimate food intake for each participant. The average of total sleep duration was 7.07 h, average of wake after sleep onset was 0.43 h, average of sleep latency was 0.14 h, and finally, average of sleep efficacy was 93.66%. Moreover, based on principal component analysis, six nutrient intake patterns were identified: the first and second patterns accounting for 53.88% of the total variance and the third and fourth patterns made up 13.6% of the total variance. Totally, the six patterns constitute 74.8% of the total variance. Our results showed that the second nutrient pattern had a negative correlation with total sleep time (P = 0.03); it was positively correlated with sleep latency (P = 0.004). The sixth nutrient pattern was negatively associated with total sleep time (P = 0.007). It was observed that higher intake of the fourth pattern had a negative correlation with total sleep time (P = 0.03). Higher intake of the fifth pattern was positively associated with sleep latency (P = 0.05). In summary, we found that nutrient patterns are correlated with sleep pattern.

  相似文献   
880.

In the present study, halophilic bacteria communities were explored in saline soils of Howze-Soltan playa in Iran with special attention to their biological activity against an aflatoxigenic Aspergillus parasiticus NRRL 2999. Halophilic bacteria were isolated from a total of 20 saline soils using specific culture media and identified by 16S rRNA sequencing in neighbor-joining tree analysis. Antifungal and antiaflatoxigenic activities of the bacteria were screened by a nor-mutant A. parasiticus NRRL 2999 using visual agar plate assay and confirmed by high-performance liquid chromatography. Among a total of 177 halophilic bacteria belonging to 11 genera, 121 isolates (68.3%) inhibited A. parasiticus growth and/or aflatoxin production. The most potent inhibitory bacteria of the genera Bacillus, Paenibacillus and Staphylococcus were distributed in three main phylogenetic clusters as evidenced by 16S rRNA sequence analysis. A. parasiticus growth was inhibited by 0.7–92.7%, while AFB1 and AFG1 productions were suppressed by 15.1–98.9 and 57.0–99.6%, respectively. Taken together, halophilic bacteria identified in this study may be considered as potential sources of novel bioactive metabolites as well as promising candidates to develop new biocontrol agents for managing toxigenic fungi growth and subsequent aflatoxin contamination of food and feed in practice.

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