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61.
Pihlavisto P Reenilä I 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2002,781(1-2):359-372
Catechol O-methyltransferase (COMT) transfers a methyl group from S-adenosyl-L-methionine to the catechol substrate in the presence of magnesium. After the characterisation of COMT more than four decades ago, a wide variety of COMT enzyme assays have been introduced. COMT activity analysis usually consists of the handling of the sample and incubation followed by separation and detection of the reaction products. Several of these assays are validated, reliable and sensitive. Besides the studies of the basic properties of COMT, the activity assay has also been applied to explore the relation of COMT to various disease states or disorders. In addition, COMT activity analysis has been applied clinically since COMT inhibitors have been introduced as adjuvant drugs in the treatment of Parkinson's disease. 相似文献
62.
Monitoring the effects of drug treatment in rat models of disease by serum protein analysis 总被引:5,自引:0,他引:5
Gianazza E Eberini I Villa P Fratelli M Pinna C Wait R Gemeiner M Miller I 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2002,771(1-2):107-130
In this review we list from literature investigations on rat serum proteins using electrophoretic techniques in connection with drug testing. From our own research work, we provide annotated two-dimensional maps of rat serum proteins under control and experimental conditions. Emphasis is on species-specific components and on the effects of acute and chronic inflammation. We discuss our project of structural proteomics on rat serum as a minimally invasive approach to pharmacological investigation, and we outline a typical experimental plan for drug testing according to the above guidelines. We then report in detail on the results of our trials of anti-inflammatory drugs on adjuvant arthritis, an animal model of disease resembling in many aspects human rheumatoid arthritis. We demonstrate a correlation between biochemical parameters and therapeutic findings and outline the advantages of the chosen methodological approach, which proved also sensitive in revealing "side effects" of the test drugs. In an appendix we describe our experimental protocol when performing two-dimensional electrophoresis of rat serum. 相似文献
63.
64.
Electrochemical biosensors have found wide application in food and clinical areas, as well as in environmental field. A large number of articles focused on horseradish peroxidase (HRP)-based biosensors have been published in the last decade, due to the capability of HRP to quantitatively detect the presence of hydrogen peroxide produced in a reaction. At present a large body of multi-enzymatic amperometric biosensors are realized by entrapping HRP together with other enzymes into a polymeric matrix; these systems represent a promising example of simple, low-cost electrochemical tools for the analysis of bioanalytes in solution, such as glucose, choline and cholesterol. Due to the fact that polymers used for HRP entrapping are soluble in organic solvents and that many solvents are strong denaturants of aquo-soluble proteins, in this paper we investigate (in particular, by circular dichroism and electron paramagnetic spectroscopies) the effect of dimethyl sulfoxide, one of the organic solvents employed for polymer solubilization, on the structure and the functionality of HRP, in order to determine the effect induced by the solvent concentration on the structure and activity of the hemoprotein. This is relevant for basic and applied biochemistry, HRP being largely employed in bioinorganic chemistry and sensor area. 相似文献
65.
Sigrell JA Eklund P Galin M Hedkvist L Liljedahl P Johansson CM Pless T Torstenson K 《Journal of structural and functional genomics》2003,4(2-3):109-114
The capacity for high throughput purification (HTP) is essential in fields such as structural genomics where large numbers of protein samples are routinely characterized in, for example, studies of structural determination, functionality and drug development. Proteins required for such analysis must be pure and homogenous and available in relatively large amounts. ÄKTAT 3D system is a powerful automated protein purification system, which minimizes preparation, run-time and repetitive manual tasks. It has the capacity to purify up to 6 different His6- or GST-tagged proteins per day and can produce 1–50 mg protein per run at >90% purity. The success of automated protein purification increases with careful experimental planning. Protocol, columns and buffers need to be chosen with the final application area for the purified protein in mind. 相似文献
66.
Fratelli M Demol H Puype M Casagrande S Villa P Eberini I Vandekerckhove J Gianazza E Ghezzi P 《Proteomics》2003,3(7):1154-1161
Protein glutathionylation is a post-translational modification consisting of the formation of a mixed disulfide between protein cysteines and glutathione (GSH). To identify proteins undergoing glutathionylation in primary rat hepatocytes and in human HepG2 hepatoma cells, we radiolabeled the intracellular GSH pool with L-[(35)S] cysteine. Cells were then exposed to oxidative stress. Proteins were separated by two-dimensional gel electrophoresis under nonreducing conditions, and glutathionylated proteins were located by autoradiography and identified by mass spectrometry after tryptic digestion. Several proteins previously not known to undergo glutathionylation were thus recognized. Among the identified proteins some are the same or belong to the same functional class as those we have already identified in a previous paper on T cell blasts (actin, nucleophosmin, phosphogluconolactonase, myosin, profilin, cyclophilin A, stress 70 protein, ubiquitin in HepG2 cells and actin, peroxiredoxin 5, cytochrome C oxidase, heat shock cognate 70 in hepatocytes) while others are newly recognized (Ran specific GTPase activating protein, histidine triad nucleotide binding protein 2 in HepG2 cells and enoyl CoA hydratase in hepatocytes). The technique described proved equally applicable to a variety of cell types. 相似文献
67.
Pia Mutikainen Lynda F. Delph 《Evolution; international journal of organic evolution》1998,52(6):1572-1582
If inbreeding depression is caused by deleterious recessive alleles, as suggested by the partial dominance hypothesis, a negative correlation between inbreeding and inbreeding depression is predicted. This hypothesis has been tested several times by comparisons of closely related species or comparisons of populations of the same species with different histories of inbreeding. However, if one is interested in whether this relationship contributes to mating-system evolution, which occurs within populations, comparisons among families within a population are needed; that is, inbreeding depression among individuals with genetically based differences in their rate of selfing should be compared. In gynodioecious species with self-compatible hermaphrodites, hermaphrodites will have a greater history of potential inbreeding via both selfing and biparental inbreeding as compared to females and may therefore express a lower level of inbreeding depression. We estimated the inbreeding depression of female and hermaphrodite lineages in gynodioecious Lobelia siphilitica in a greenhouse experiment by comparing the performance of selfed and outcrossed progeny, as well as sibling crosses and crosses among subpopulations. We did not find support for lower inbreeding depression in hermaphrodite lineages. Multiplicative inbreeding depression (based on seed germination, juvenile survival, survival to flowering, and flower production in the first growing season) was not significantly different between hermaphrodite lineages (δ = 0.30 ± 0.08) and female lineages (δ = 0.15 ± 0.18), although the trend was for higher inbreeding depression in the hermaphrodite lineages. The population-level estimate of inbreeding depression was relatively low for a gynodioecious species (δ = 0.25) and there was no significant inbreeding depression following biparental inbreeding (δ = 0.01). All measured traits showed significant variation among families, and there was a significant interaction between family and pollination treatment for four traits (germination date, date of first flowering, number of flowers, and aboveground biomass). Our results suggest that the families responded differently to selfing and outcrossing: Some families exhibited lower fitness following selfing whereas others seemed to benefit from selfing as compared to outcrossing. Our results support recent simulation results in that prior inbreeding of the lineages did not determine the level of inbreeding depression. These results also emphasize the importance of determining family-level estimates of inbreeding depression, relative to population-level estimates, for studies of mating-system evolution. 相似文献
68.
Christian Schobert Lucian Baker Judit Szederkényi Pia Großmann Ewald Komor Hiroaki Hayashi Mitsuo Chino William J. Lucas 《Planta》1998,206(2):245-252
The mature, functional sieve-tube system in higher plants is dependent upon protein import from the companion cells to maintain
a functional long-distance transport system. Soluble proteins present within the sieve-tube lumen were investigated by analysis
of sieve-tube exudates which revealed the presence of distinct sets of polypeptides in seven monocotyledonous and dicotyledonous
plant species. Antibodies directed against sieve-tube exudate proteins from Ricinus communis L. demonstrated the presence of shared antigens in the phloem sap collected from Triticum aestivum L., Oryza sativa L., Yucca filamentosa L., Cucurbita maxima Duch., Robinia pseudoacacia L. and Tilia platyphyllos L. Specific antibodies were employed to identify major polypeptides. Molecular chaperones related to Rubisco-subunit-binding
protein and cyclophilin, as well as ubiquitin and the redox proteins, thioredoxin h and glutaredoxin, were detected in the
sieve-tube exudate of all species examined. Actin and profilin, a modulator of actin polymerization, were also present in
all analyzed phloem exudates. However, some proteins were highly species-specific, e.g. cystatin, a protease-inhibitor was
present in R. communis but was not detected in exudates from other species, and orthologs of the well-known squash phloem lectin, phloem protein
2, were only identified in the sieve-tube exudate of R. communis and R. pseudoacacia. These findings are discussed in terms of the likely roles played by phloem proteins in the maintenance and function of the
enucleate sieve-tube system of higher plants.
Received: 12 February 1998 / Accepted: 16 March 1998 相似文献
69.
70.
Maria Pia Bucci Sylvette Wiener-Vacher Clémence Trousson Olivier Baud Valerie Biran 《PloS one》2015,10(3)