Measurement of the spin concentration of metalloprotein samples from saturation-magnetization data with particular reference to cytochrome c oxidase.
Measurement of the spin concentration of metalloprotein samples from saturation-magnetization data with particular reference to cytochrome c oxidase.
复制标题
根据饱和磁化数据测量金属蛋白样品的自旋浓度，特别参考细胞色素 c 氧化酶。
DOI：
10.1042/bj3050871
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Wilson,MT
中科院分区：
文献类型：
--
作者：
Peterson,J;Day,EP;Pearce,LL;Wilson,MT
一种利用超导量子干涉装置（SQUID）磁强计从金属蛋白样品中获得高质量饱和磁化数据的方案已经被报道[E]。P. Day， T. A. Kent， P. A. Lindahl， E. m<e:1> nck， W. H. Orme-Johnson， H. Roder和A. Roy（1987）《生物物理学》[j].化学工程学报，1997,18(2):437-463。在研究了几十种不同的金属蛋白衍生物之后，该方法得到了进一步的改进，特别是在样品制备领域。描述了目前使用的样品处理程序的细节，此外，还考虑了验证顺磁杂质污染仍然微不足道的关键问题。重要的是，它表明，一个独立的确定数量的顺磁样品存在于磁力计是不希望的。如果收集到足够的饱和磁化数据，可以在随后的拟合过程中确定自旋浓度，则可以获得有关所研究系统的基态磁性能的更可靠的参数。通过对辣根过氧化物酶的铁细胞色素c、铁细胞色素c和苯甲羟肟酸加合物的磁化研究，证明了该方法的有效性。饱和磁化测量常规确定自旋浓度的能力在接受值的+/- 4%以内是牢固建立的。此外，对细胞色素c氧化酶的静息和完全还原衍生物进行了饱和磁化研究。这些结果说明了该技术在探测一些被证明难以用其他方法研究的系统方面的有用性。从这些细胞色素c氧化酶和其他整数自旋系统中获得有意义的数据所固有的困难增加了。
A protocol for obtaining high-quality saturation-magnetization data from metalloprotein samples, employing a superconducting quantum interference device (SQUID) magnetometer, has previously been reported [E. P. Day, T. A. Kent, P. A. Lindahl, E. Münck, W. H. Orme-Johnson, H. Roder and A. Roy (1987) Biophys. J. 52, 837-853 and E. P. Day (1993) Methods Enzymol. 227, 437-463]. Following studies of several dozen different metalloprotein derivatives, the methodology has been further refined, particularly in the area of sample preparation. The details of the sample-handling procedures now in use are described, and moreover, the critical issue of verifying that contamination by paramagnetic impurities remains insignificant is considered. Importantly, it is shown that an independent determination of the quantity of paramagnetic sample present in the magnetometer is undesirable. Much more reliable parameters concerning the ground-state magnetic properties of the system under study are obtained if enough saturation-magnetization data are collected to enable the spin concentration to be determined during the subsequent fitting procedure. As proof of the validity of this method, the results of magnetization studies on ferricytochrome c, ferrocytochrome c and the benzohydroxamic acid adduct of horseradish peroxidase are presented. The ability of saturation-magnetization measurements to routinely determine spin concentration to within +/- 4% of accepted values is firmly established. In addition, a saturation-magnetization study has been performed on resting and fully reduced derivatives of cytochrome c oxidase. These results provide an illustration of the usefulness of the technique in probing some systems which have proved difficult to study by other methods. The increased difficulties inherent in obtaining meaningful data from these cytochrome c oxidase and other integer spin systems are delineated.