Biological pretreatment of lignocellulosic biomass by fungi can represent a low-cost and eco-friendly alternative to physicochemical methods to facilitate enzymatic hydrolysis. However, fungal metabolism can cause cellulose loss and it is therefore necessary to use the appropriate fungal strain-biomass type combination. In this work, the effects of biological pretreatments carried out by five different fungi on enzymatic hydrolysis of wheat straw were investigated. The best results were obtained with a Ceriporiopsis subvermispora strain, which minimized weight and cellulose losses and gave the highest net sugar yield (calculated with respect to the holocellulose content of the untreated straw), up to 44 % after a 10-week pretreatment, more than doubling the yields obtained with the other isolates. Moreover, prolonging the pretreatment from 4 up to 10 weeks produced a 2-fold increase, up to 60 %, in digestibility (sugar yield, calculated considering the holocellulose content of the pretreated material). The hemicellulose content of the pretreated material resulted inversely correlated with digestibility, and it could thus be utilized as an index of the pretreatment efficacy. Finally, a correlation was also found between digestibility and the difference between the absorbance values at 290 and 320 nm of pretreated wheat straw extracts.
The sodium iodide symporter (NIS) mediates iodide (I(-)) transport in the thyroid gland and other tissues and is of increasing importance as a therapeutic target and nuclear imaging reporter. NIS activity in vitro is currently measured with radiotracers and electrophysiological techniques. We report on the development of a novel live cell imaging assay of NIS activity using the I(-)-sensitive and genetically encodable yellow fluorescent protein (YFP) variant YFP-H148Q/I152L. In FRTL-5 thyrocytes stably expressing YFP-H148Q/I152L, I(-) induced a rapid and reversible decrease in cellular fluorescence characterized by 1) high affinity for extracellular I(-) (35 muM), 2) inhibition by the NIS inhibitor perchlorate, 3) extracellular Na(+) dependence, and 4) TSH dependence, suggesting that fluorescence changes are due to I(-) influx via NIS. Individual cells within a population of FRTL-5 cells exhibited a 3.5-fold variation in the rate of NIS-mediated I(-) influx, illustrating the utility of YFP-H148Q/I152L to detect cell-to-cell difference in NIS activity. I(-) also caused a perchlorate-sensitive decrease in YFP-H148Q/I152L fluorescence in COS-7 cells expressing NIS but not in cells lacking NIS. These results demonstrate that YFP-H148Q/I152L is a sensitive biosensor of NIS-mediated I(-) uptake in thyroid cells and in nonthyroidal cells following gene transfer and suggest that fluorescence detection of cellular I(-) may be a useful tool by which to study the pathophysiology and pharmacology of NIS.
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