ELECTRON ELECTRON DIPOLAR DISTANCE MEASUREMENTS IN DISORDERED SOLIDS

利用EPR波谱对氮氧自由基自旋进行择位 最近， 作者：Shin，Beth and Hubbell 我们已经研究了 利用高频EPR提取双自由基之间的距离 氮氧自旋标记。 我们研究中使用的4K-li螺旋肽 是基于丙氨酸的肽，其中引入了两个半胱氨酸 在肽序列的不同位置。 的半胱氨酸残基 然后用甲硫基磺酸盐自旋标记物（MTSSL）标记。 对各种双自旋的高频谱进行了测量 标记的肽; AK 6L是单标记的肽，AK 417是标记的肽， 在位置（i，i+13）处，AK 49在位置（i，i+S）处，AK 68在位置（i，i +13）处， 在位置（i，i+2）处的AK 69、在位置（i，i+3）处的AK 69和在位置（i，i+4）处的AK 48。 螺旋肽每圈有3.5个残基， 因此，对氮氧EPR谱的影响最大， 肽AK 48（i，i+4）和AK 69（i，i+3），其中两个自旋标记是 非常接近（7-9 A间隔）。 分析了增宽和 高频EPR共振的分裂使得 确定两个氮氧自旋标记之间的距离。 的 EPR共振的加宽/分裂在以下方面最明显： EPR粉末高场边缘的~ 4 N超精细共振 格局 高频CW EPR谱对两种信号都很敏感， 偶极距和电子的相对取向 g张量和电子偶极张量。

The use of EPR spectroscopy of site-selective nitroxide spin labeled proteins for measuring distances has recently been pioneered by Shin, Beth and Hubbell. We have investigated the advantages of using high frequency EPR to extract distances between biradical nitroxide spin labels. The 4K-li helical peptide used in our studies is an alanine based peptide where two cysteines have been introduced at different positions in the peptide sequence. The cysteine residues are then labeled with the methanethiosulfonate spin label (MTSSL). High frequency spectra have been obtained for a variety of doubly spin labeled peptides; AK6L is the singly labeled peptide, AK417 is labeled at positions (i,i+13), AK49 at positions (i,i+S), AK68 at positions (i,i+2), AK69 at positions (i,i+3), and AK48 at positions (i,i+4). There are 3.5 residues/turn of the helical peptide and one sees, therefore, the greatest effect on the nitroxide EPR spectrum for peptides AK48 (i,i+4) and AK69 (i,i+3), where the two spin labels are in close proximity (7-9 A separation). Analysis of the broadening and splitting of the high frequency EPR resonances enables the determination of the distance between the two nitroxide spin labels. The broadening/splitting of the EPR resonances is most pronounced on the '4N hyperfine resonances on the high field edge of the EPR powder pattern. The high frequency CW EPR spectra are sensitive to both the dipolar distance and the relative orientation of the electron g-tensors and the electron dipolar tensor.