Borkman R F, Phillips S R
Exp Eye Res. 1985 Jun;40(6):819-26. doi: 10.1016/0014-4835(85)90127-7.
The efficiency of electronic energy transfer from tyrosine to tryptophan residues in the lens protein calf gamma II crystallin has been determined from measurements of fluorescence excitation spectra at 25 degrees C and independently from phosphorescence spectroscopy at 77 K. The total transfer efficiency from the fifteen tyrosines to the four tryptophans in native calf gamma II was found to be t = 78 +/- 10%, at 25 degrees C. The expected value based on Förster theory and the X-ray structure of the protein is t = 83%, in good agreement with the present experimental result. The transfer efficiency measured in denatured calf gamma II at 25 degrees C was t = 20 +/- 10%, in good agreement with the value t = 25% expected for the completely denatured protein, based on Förster theory and the known amino acid sequence. These results are of interest for several reasons. First, energy transfer can provide a simple , experimental confirmation of lens protein structures determined from X-ray data or from computer modeling studies. Second, the present studies show that energy transfer measurements can be used to monitor the effect of denaturants on lens protein structure, an aspect not readily investigated by X-ray crystallography. Third, the present electronic energy transfer studies may be relevant to understanding the mechanism of UV photodamage in lens proteins and hence in whole lenses.
通过在25℃下测量荧光激发光谱以及独立地在77K下进行磷光光谱测量，已确定了晶状体蛋白小牛γII晶状体蛋白中从酪氨酸到色氨酸残基的电子能量转移效率。在25℃下，天然小牛γII中从15个酪氨酸到4个色氨酸的总转移效率为t = 78±10%。基于Förster理论和该蛋白质的X射线结构的预期值为t = 83%，与目前的实验结果吻合良好。在25℃下变性小牛γII中测得的转移效率为t = 20±10%，与基于Förster理论和已知氨基酸序列对完全变性蛋白质预期的t = 25%的值吻合良好。这些结果因几个原因而备受关注。首先，能量转移可以为从X射线数据或计算机建模研究确定的晶状体蛋白结构提供一个简单的实验验证。其次，目前的研究表明，能量转移测量可用于监测变性剂对晶状体蛋白结构的影响，这是X射线晶体学不易研究的一个方面。第三，目前的电子能量转移研究可能与理解晶状体蛋白以及整个晶状体中紫外线光损伤的机制相关。
