The experimental determination of the binding constant of a drug for its target molecule is of considerable importance. It is a basic experimental parameter in a variety of studies, such as the prediction of drug efficiency, or in the pharmacokinetic drug interaction. DNA-binding drugs have been reported to be able to interfere in a sequence dependent manner with biological functions such as topoisomerase activity, restriction of enzyme cleavage of DNA, protein-DNA interactions and the activity of transcription factors, leading to alteration of gene expression. This effect could have important practical application in the experimental therapy of human pathologies, including neoplastic diseases and viral, or microbial infections. The assessment of the biological activity of DNA-binding drugs by polymerase chain reaction, footprinting, gel retardation and in vitro transcription studies was recently reported. However, most of these techniques are steady-state methodologies and therefore are not suitable for an easy determination of the binding activity of DNA-binding drugs to target DNA and the stability of drugs-DNA complexes. Direct real-time observation and measurement of the interaction between DNA-binding drug and target DNA sequence is a subject of interest for drug discovery and development. The recent development of biosensors, based on surface plasmon resonance (SPR) technology, enables monitoring of a variety of biospecific interactions of DNA-binding drugs with target DNA elements in real-time. The present review is designed to indicate the theoretical background of SPR-based biosensor technology as well as to present the great variety of measurements and modes of interaction kinetics that can be performed with these techniques. In addition, some of the most recent studies in determining the binding constant and stoichiometry of DNA-binding drugs to target DNA with SPR technology are reviewed and the available theoretical aspects necessary for the comprehension of the experiments are provided.
The optimum concentrations of nutrients (glucose and yeast extract) and cultivation conditions (concentration of sodium chloride, pH and incubation time) on docosahexaenoic acid (DHA) production by Schizochytrium sp. S31 in flasks at 30C were studied. Experiment design employed fractional factorial design, path of steepest ascent, central composite design and response surface methodology (RSM). The empirical model developed by RSM was adequate to describe the relationships between the studied factors and the response of DHA production. Based on contour plots and canonical analysis, the optimal conditions for maximizing DHA production (516 mg/L) were at 27.98 g glucose/L, 4.52 g yeast extraction/L, 24.82 g sodium chloride/L, pH 6.96 and incubation for 4 days at 30C. Experimental verification of the optimal conditions resulted in about 97% of the predicted DHA production by the model.
The symbiotic roles of rhizobia with legumes have been extensively made use of in the agricultural field. An unresolved question about the infection process is the nature of the Rhizobium enzymes that degrade the plant cell wall. For the bacteria in the plant root-hair wall, carboxymethylcellulase (EC 3.2.1.4) could be one of the key enzymes in the early symbiotic process. Electron microscopic immunogold labeling experiments were performed with ultrathin sections of Sinorhizobium fredii CCRC15769. The results preliminarily indicated that antigenic determinants of membrane-bound carboxymethylcellulase are exposed, at the outside of the cytoplasmic membrane, to the periplasmic space; immuno-gold particles with 10nm diameter were hardly ever observed in the cytoplasm and other spaces.
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