Davis D J
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville 72701.
Arch Biochem Biophys. 1990 Jan;276(1):1-5. doi: 10.1016/0003-9861(90)90001-f.
The tryptophan fluorescence properties of the flavoprotein ferredoxin:NADP reductase have been examined. Although not sensitive to changes in pH or salt concentration, the tryptophan fluorescence is affected by the presence of substrates for the flavoprotein. While NADP addition results in a slight quenching of the fluorescence, ferredoxin decreases the fluorescence by nearly 50%, suggesting the presence of tryptophan in or near the ferredoxin binding site. Titration of this effect gives a dissociation constant for the ferredoxin: flavoprotein complex which is similar to that obtained by spectral perturbations. This approach has also been used to demonstrate that a chemically modified ferredoxin which does not produce spectral perturbations when added to flavoprotein is capable of interacting with the flavoprotein although with a higher dissociation constant than for native ferredoxin.
NADP还原酶的色氨酸荧光特性进行了研究。尽管色氨酸荧光对pH或盐浓度的变化不敏感，但它会受到黄素蛋白底物存在的影响。添加NADP会导致荧光略有猝灭，而铁氧化还原蛋白则会使荧光降低近50%，这表明在铁氧化还原蛋白结合位点或其附近存在色氨酸。对这种效应进行滴定得到铁氧化还原蛋白：黄素蛋白复合物的解离常数，该常数与通过光谱扰动获得的解离常数相似。这种方法还被用于证明，一种化学修饰的铁氧化还原蛋白，当添加到黄素蛋白中时不会产生光谱扰动，但它能够与黄素蛋白相互作用，尽管其解离常数比天然铁氧化还原蛋白的解离常数更高。
