Barboy N, Feitelson J
Biochemistry. 1978 Nov 14;17(23):4923-6. doi: 10.1021/bi00616a010.
N-Acetyltryptophanamide (NATA), when illuminated anywhere within the 280-nm absorption band, has an emission lifetime of 3.1 ns. The tryptophan residues in liver alcohol dehydrogenase (LADH), however, when excited at 280 nm exhibit two lifetimes of r1 = 2.2 and of r2 = 5.7 ns. Excitation at 300 nm yields a single decay of 5.0 ns. It is shown that at the latter wavelength, only the two (equivalent) tryptophan residues buried within the LADH structure are excited. The reaction rate of the NATA fluorescence quenching by ionic and nonionic quenchers is practically independent of the temperature (between 5 and 41 degrees C). The same substances were used to quench the tryptophan fluorescence in LADH. Here (in the same temperature range), the quenching rate decreases drastically with a decrease in temperature. These findings are discussed in terms of conformational fluctuations in LADH, whereby the temporal movement of the polypeptide chains opens channels through which the above quencher molecules can diffuse and reach the tryptophan residues located within the enzyme structure.
N-乙酰色氨酸酰胺（NATA）在280纳米吸收带内的任何位置受到光照时，发射寿命为3.1纳秒。然而，肝脏乙醇脱氢酶（LADH）中的色氨酸残基在280纳米处激发时，呈现出两种寿命，τ1 = 2.2纳秒和τ2 = 5.7纳秒。在300纳米处激发产生5.0纳秒的单一衰减。结果表明，在后者波长下，只有埋在LADH结构内的两个（等效）色氨酸残基被激发。离子型和非离子型猝灭剂对NATA荧光的猝灭反应速率实际上与温度（5至41摄氏度之间）无关。同样的物质被用于猝灭LADH中的色氨酸荧光。在此（在相同温度范围内），猝灭速率随温度降低而急剧下降。这些发现根据LADH中的构象波动进行了讨论，即多肽链的瞬时运动打开了通道，上述猝灭剂分子可以通过这些通道扩散并到达位于酶结构内的色氨酸残基。
