The tautomeric composition of a solution of ammonium 3-deoxy-d-manno-octulosonate (KDO, 1a) in D2O at 28° was assessed by means of 13 C-F.t.-n.m.r. spectroscopy. The results revealed the presence of 6?0 and 11 % of the α and β anomers of the pyranose, and 20 and 9 % of the two furanoses, and suggested, but did not unequivocally prove, that the major furanose form is the α anomer. To facilitate interpretation of the spectral results for 1, ammonium 3,5-dideoxy-d-arabino(or ribo)-octulosonate (3a) was prepared by the reaction of 5-deoxy-d-erythro-pentose with sodium oxalacetate at pH 11. A chromatographically homogeneous, noncrystalline sample of 3 was obtained by lyophilization, and characterized as its (4-nitrophenyl)hydrazone (m.p. 162-163°). The 13C-n.m.r. spectrum of a solution of 3a in D2O revealed it to be substantially all in the α-pyranose form. No signals were obtained for the possible 1,4-lactone of 3. As the 1,5-lactone and furanose forms are impossible for 3, it exhibited no signals analogous to those attributed to furanoid 1. On the basis of these results for 3, the two lactone forms of 1 were excluded from consideration, and the three pairs of 13C-n.m.r. signals observed at ≈45, 86, and 104 p.p.m. were assigned to the furanose forms of 1. 相似文献
Probiotics and Antimicrobial Proteins - In this study, a bacterial strain COFCAU_P1, isolated from the digestive tract of a freshwater teleost rohu (Labeo rohita), was identified as Bacillus... 相似文献
Use of probiotics as the biocontrol agent for disease prevention in aquaculture is gaining importance as an alternative to the indiscriminate use of antibiotics and other chemotherapeutics. In view of this trend, the probiotic properties of a potent antagonistic bacterium, Pseudomonas aeruginosa FARP72, was characterized in terms of safety, antagonistic activities, in vitro immunomodulation, and in vivo disease resistance. Immunomodulatory activity was ascertained by measuring the production of intracellular superoxide anion, nitric oxide, total leukocyte peroxidase content, and the leukocyte proliferation in head kidney leukocytes. The bacterium isolated from the skin mucus of freshwater catfish Clarias batrachus was harmless to Labeo rohita. It showed inhibitory activity against Aeromonas caviae, A. hydrophila, Edwardsiella tarda, Pseudomonas putida, and Streptococcus agalactiae as revealed by cross and parallel streaking methods. Significantly higher superoxide anion and nitric oxide production, peroxidase content, and proliferative responses of leucocytes delineated the strains’ ability to interact with immune cells to activate the immune system in vitro. Significant growth inhibition of A. hydrophila from 1.55 × 105 CFU/mL was observed when co-cultured with P. aeruginosa FARP72 in phosphate-buffered saline (PBS) at levels ranging from 2.61 × 107 to 2.61 × 109 CFU/mL in 10 days. P. aeruginosa FARP72 increased the survival rate of rohu fingerlings against pathogenic A. hydrophila challenge in biocontrol study in vivo as determined by cohabitation challenge. These results suggest that P. aeruginosa FARP72 is a potential probiotic strain and can be used in aquaculture to improve the health status and disease resistance of fish.
We have cloned and overexpressed multidrug transporter CaMdr1p as a green fluorescent protein-tagged protein to show its capability to extrude drug substrates. The drug extrusion was sensitive to pH and energy inhibitors and displayed selective substrate specificity. CaMdr1p has a unique and conserved antiporter motif, also called motif C [G(X6)G(X3)GP(X2)GP(X2)G], in its transmembrane segment 5 (TMS 5). Alanine scanning of all the amino acids of the TMS 5 by site-directed mutagenesis highlighted the importance of the motif, as well as that of other residues of TMS 5, in drug transport. The mutant variants of TMS 5 were placed in four different categories. The first category had four residues, G244, G251, G255, and G259, which are part of the conserved motif C, and their substitution with alanine resulted in increased sensitivity to drugs and displayed impaired efflux of drugs. Interestingly, first category mutants, when replaced with leucine, resulted in more dramatic loss of drug resistance and efflux. Notwithstanding the location in the core motif, the second category included residues which are part of the motif, such as P260, and those which were not part of the motif, such as L245, W248, P256, and F262, whose substitution with alanine resulted in a severe loss of drug resistance and efflux. The third category included G263, which is a part of motif C, but unlike other conserved glycines, its replacement with alanine or leucine showed no change in the phenotype. The replacement of the remaining 11 residues of the fourth category did not result in any change. The putative helical wheel projection showed clustering of functionally critical residues to one side and thus suggests an asymmetric nature of TMS 5. 相似文献
Protoporphyrin IX and its derivatives are used as photosensitizers in the photodynamic therapy of cancer. Protoporphyrin IX
penetrates into human red blood cells and releases oxygen from them. This leads to a change in the morphology of the cells.
Spectrophotometric studies reveal that protoporphyrin IX interacts with haemoglobin and myoglobin forming ground state complexes.
For both proteins, the binding affinity constant decreases, while the possible number of binding sites increases, as the aggregation
state of the porphyrin is increased. The interactions lead to conformational changes of both haemoglobin and myoglobin as
observed in circular dichroism studies. Upon binding with the proteins, protoporphyrin IX releases the heme-bound oxygen from
the oxyproteins, which is dependent on the stoichiometric ratios of the porphyrin: protein. The peroxidase activities of haemoglobin
and myoglobin are potentiated by the protein-porphyrin complexation. Possible mechanisms underlying the relation between the
porphyrin-induced structural modifications of the heme proteins and alterations in their functional properties have been discussed.
The findings may have a role in establishing efficacy of therapeutic uses of porphyrins as well as in elucidating their mechanisms
of action as therapeutic agents. 相似文献
COPII vesicles assemble at ER subdomains called transitional ER (tER) sites, but the mechanism that generates tER sites is unknown. To study tER biogenesis, we analyzed the transmembrane protein Sec12, which initiates COPII vesicle formation. Sec12 is concentrated at discrete tER sites in the budding yeast Pichia pastoris. We find that P. pastoris Sec12 exchanges rapidly between tER sites and the general ER. The tER localization of Sec12 is saturable and is mediated by interaction of the Sec12 cytosolic domain with a partner component. This interaction apparently requires oligomerization of the Sec12 lumenal domain. Redistribution of P. pastoris Sec12 to the general ER does not perturb the localization of downstream tER components, suggesting that Sec12 and other COPII proteins associate with a tER scaffold. These results provide evidence that tER sites form by a network of dynamic associations at the cytosolic face of the ER. 相似文献
Extensive investigation into plant response and adaptation to diverse osmotic stresses like high salt/dehydration/low temperature,
involving a broad spectrum of cellular physiological and biochemical changes, is essential to unravel intrinsic mechanism
to mitigate against such stresses. In our previous communications, we conducted biochemical analyses of indica rice varieties,
subjected to exogenous salt/abscisic acid-mediated oxidative stress. The aim of this study was to compare differential biochemical
responses of the salt-sensitive (IR-29), salt-tolerant (Pokkali) and aromatic (Pusa Basmati or PB) rice varieties during polyethylene
glycol (PEG)-induced dehydration stress. The greater susceptibility of IR-29 and PB, to water scarcity, was reflected by the
higher toxic Na+ and putrescine accumulation, considerable decrease in (reduced/oxidized) glutathione, maximal increment in protease activity
and greater downregulation of nitrate reductase activity. On the other hand, Pokkali appeared to suffer lesser damages as
evidenced from much lower endogenous Na+ but higher K+, Ca2+ and Mg2+ accumulation, registering the highest levels of osmolytes like glycinebetaine and higher polyamines (spermidine and spermine)
accounting to improved relative water content, higher (reduced/oxidized) glutathione, maximal induction of the enzyme phenylalanine
ammonia-lyase and practically unhindered nitrate reductase activity, following PEG treatment. The highest induction of sugars
and proline in IR-29 and PB probably played the osmoprotective/antioxidative functions, enabling to a certain extent to heighten
their lipoxygenase inhibition or H2O2 scavenging potential, more than Pokkali, to ward off oxidative damages and sustain survival under critical dehydrated situations.
Thus, the salt-tolerant Pokkali also showed prominent dehydration-tolerance properties, whereas the aromatic rice PB, almost
identical in their biochemical responses to IR-29, showed greater sensitivity to PEG-mediated water deficit. 相似文献
In a hydroponic setting, we investigated the possible role of phytochelatins (metal-binding peptides) in the lead (Pb) tolerance of vetiver grass (Vetiveria zizanioides L.). Pb was added to the nutrient medium at concentrations ranging from 0 to 1,200 mg L?1. Furthermore, we simulated the effect of soil phosphorus (P) on potentially plant available Pb by culturing vetiver grass in P-rich nutrient media. After 7 days of exposure to Pb, we evaluated the Pb uptake by vetiver grass. Results indicate that vetiver can accumulate Pb up to 3,000 mg kg?1 dry weight in roots with no toxicity. Formation of lead phosphate inhibited Pb uptake by vetiver, suggesting the need for an environmentally safe chelating agent in conjunction with phytoremediation to clean up soils contaminated with lead-based paint. Unambiguous characterization of phytochelatins (PCn) was possible using high pressure liquid chromatography coupled with electrospray ionization mass spectrometry (LC-ESMS). Vetiver shows qualitative and quantitative differences in PCn synthesis between root and shoot. In root tissue from vetiver exposed to 1,200 mg Pb L-1, phytochelatins ranged from PC1 to PC3. Collision-induced dissociation of the parent ion allowed confirmation of each PCn based on the amino acid sequence. Possible Pb-PC1 and Pb2-PC1 complexes were reported in vetiver root at the highest Pb concentration. The data from these experiments show that the most probable mechanism for Pb detoxification in vetiver is by synthesizing PCn and forming Pb–PCn complexes. 相似文献