MAGNETIC SUSCEPTIBILITY STUDIES OF METALLOPROTEINS

金属蛋白的磁化率研究具有很大的潜在意义 价值。很少有其他技术可以测量整数的磁性 自旋态和其他顺磁态的EPR无声 松弛扩大等原因。 金属蛋白敏感性研究的困难在于 由于磁感计缺乏灵敏度，因此需要高度 浓缩样品(大于1000微米)。浓缩度 光学或EPR表征之后的程序通常会导致更改 在样品的氧化还原状态下。的氧化还原状态的模糊性。 合格样品，样品杂质水平的不确定度 顺磁性，以及内在缺乏解决静态 磁化率法与EPR等共振技术的比较 和穆斯堡尔谱学导致了不可靠的解释 敏感度实验。此外，高等教育的一个根本问题是 分子量蛋白质已经不可能达到 敏感性试验所需的浓度。 我们已经开发了新的技术来测量Low的磁性 浓缩(大于50微米)金属蛋白样品 商用超导鱿鱼磁感仪。我们已经将这一新的 通过开发一种电子顺磁共振和穆斯堡尔谱能力 多仪器样品保持器。我们现在可以研究同样的冰冻 金属蛋白样品(不解冻样品或改变其 浓度或持有者)在研究之前与EPR和穆斯堡尔都有联系 用感受器。通过这种技术的组合，氧化还原 可以完全定义敏感性样本的状态，并且其水平 测量的杂质顺磁性可靠地解释了 敏感性数据是可能的。此外，新的实验结合了 低温光解和磁化率测量现在 可行。 我们将研究细菌和哺乳动物细胞色素氧化酶，一种细菌 Rieske蛋白和其他有代表性的[2Fe-2S]蛋白，大肠杆菌亚硫酸盐 还原酶和铁氧还蛋白II。 我们的目标之一是为研究猪传染性支气管炎易感性奠定基础 室温下的生化动力学。

Magnetic susceptibility studies of metalloproteins are potentially of great value. Few other techniques can measure the magnetic properties of integer spin states and of other paramagnetic states which are EPR-silent for reasons such as relaxation broadening. The difficulty with susceptibility studies of metalloproteins has been the lack of sensitivity of susceptometers leading to a requirement for highly concentrated samples (greater than 1000 MuM). The concentration procedure following optical or EPR characterization often causes changes in the sample's redox state. Ambiguity in the redox state of the suceptibility sample, uncertainties in the level of the sample's impurity paramagnetism, and the intrinsic lack of resolution of the static susceptibility method when compared with resonance techniques such as EPR and Mossbauer spectroscopy have led to unreliable interpretation of susceptibility experiments. Moreover, a fundamental problem for high molecular weight proteins has been the impossibility of achieving the concentration necessary for a susceptibility experiment. We have developed new techniques to measure the magnetic properties of low concentration (greater than 50 MuM) metalloprotein samples using a commercial superconducting SQUID susceptometer. We have combined this new capability with EPR and Mossbauer spectroscopy by developing a multi-instrument sample holder. We can now study the same frozen metalloprotein sample (without thawing the sample or changing its concentration or holder) with both EPR and Mossbauer prior to its study with the susceptometer. With this combination of techniques the redox state of the susceptibility sample can be fully defined and its level of impurity paramagnetism measured making reliable interpretation of the susceptibility data possible. In addition, new experiments combining photolysis at low temperatures with susceptibility measurements are now feasible. We will study bacterial and mammalian cytochrome oxidase, a bacterial Rieske protein and other representative [2Fe-2S] proteins, E. coli sulfite reductase, and D. gigas ferredoxin II. One of our goals is to lay the ground work for susceptibility studies of biochemical kinetics at room temperature.