Excited-state lifetime studies of the three tryptophan residues in the N-lobe of human serum transferrin
Excited-state lifetime studies of the three tryptophan residues in the N-lobe of human serum transferrin
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DOI：
10.1002/pro.287
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发表时间：
2010-01-01
期刊：
影响因子：
8
通讯作者：
Jameson, David M.
中科院分区：
文献类型：
--
作者：
James, Nicholas G.;Ross, Justin A.;Jameson, David M.
通过监测Trp荧光发射和寿命的变化，研究了从人血清转铁蛋白（hTF）N叶内的位置8，128和264处的三个Trp残基到配体到金属电荷转移带的能量转移。来自hTF N-叶的荧光发射由Trp 264主导，如当不存在该Trp残基时量子产率降低82%所揭示的。荧光寿命测定的突变体含有一个或两个色氨酸残基的多频相位荧光。这些样品的衰变最好用两个或三个离散寿命或单峰洛伦兹分布来描述。含Trp 128和Trp 264的样品的离散寿命和寿命分布的中心受到铁的影响。铁去除的分布宽度变窄，并与占主导地位的荧光团，Trp 264的动态迁移率的下降是一致的。然而，当铁存在时，量子产率和寿命都较低，但不成比例。铁对量子产率的较大影响表明非激发态淬灭，即，静态猝灭这些实验的结果提供了定量数据，有力地表明福斯特共振能量转移不是在hTF的N-叶中色氨酸淬灭的唯一来源。
The energy transfer from the three Trp residues at positions 8, 128, and 264 within the human serum transferrin (hTF) N-lobe to the ligand to metal charge transfer band has been investigated by monitoring changes in Trp fluorescence emission and lifetimes. The fluorescence emission from hTF N-lobe is dominated by Trp264, as revealed by an 82% decrease in the quantum yield when this Trp residue is absent. Fluorescence lifetimes were determined by multifrequency phase fluorometry of mutants containing one or two Trp residues. Decays of these samples are best described by two or three discrete lifetimes or by a unimodal Lorentzian distribution. The discrete lifetimes and the center of the lifetime distribution for samples containing Trp128 and Trp264 are affected by iron. The distribution width narrows on iron removal and is consistent with a decrease in dynamic mobility of the dominant fluorophore, Trp264. Both the quantum yield and the lifetimes are lower when iron is present, however, not proportionally. The greater effect of iron on quantum yields is indicative of nonexcited state quenching, i.e., static quenching. The results of these experiments provide quantitative data strongly suggesting that Forster resonance energy transfer is not the sole source of Trp quenching in the N-lobe of hTF.