Busch J L, Breton J L, Bartlett B M, Armstrong F A, James R, Thomson A J
School of Chemical Sciences, University of East Anglia, Norwich NR4 7TJ, UK.
Biochem J. 1997 Apr 1;323 ( Pt 1)(Pt 1):95-102. doi: 10.1042/bj3230095.
The 8Fe ferredoxin III from Desulfovibrio africanus is a monomeric protein which contains two [4Fe-4S]2+/1+ clusters, one of which is labile and can readily and reversibly lose one Fe under oxidative conditions to yield a [3Fe-4S]1+/0 cluster. This 4Fe cluster has an S = 3/2 ground sping state insteaed of S = 1/2 in the reduced +1 state [George, Armstrong, Hatchikian and Thomson (1989) Biochem. J. 264, 275-284]. The co-ordination to this cluster is unusual in that an aspartate (Asp14, D14, is found where a cysteine residue normally occurs. Using a mutant protein obtained from the overexpression in Escherichia coli of a synthetic gene in which Asp14, the putative ligand to the removable Fe, has been changed to Cys, we have studied the cluster interconversion properties of the labile cluster. Analysis by EPR and magnetic-circular-dichroism spectroscopies showed that the Asp14 --> Cys (D14C) mutant contains two [4Fe-4S]2+/1+ clusters, both with S = 1/2 in the reduced state. Also, unlike in native 8Fe D. africanus ferredoxin III, the 4Fe <--> 3Fe cluster interconversion reaction was found to be sluggish and did not go to completion. It is inferred that the reversibility of the reaction in the native protein is due to the presence of the aspartate residue at position 14 and that this residue might protect the [3Fe-4S] cluster from further degradation.
来自非洲脱硫弧菌的8Fe铁氧化还原蛋白III是一种单体蛋白，它含有两个[4Fe-4S]2+/1+簇，其中一个不稳定，在氧化条件下能轻易且可逆地失去一个铁，生成一个[3Fe-4S]1+/0簇。这个4Fe簇具有S = 3/2的基态自旋态，而不是还原态+1时的S = 1/2 [乔治、阿姆斯特朗、哈奇基安和汤姆森（1989年）《生物化学杂志》264卷，275 - 284页]。与这个簇的配位情况不同寻常，因为在通常出现半胱氨酸残基的位置发现了一个天冬氨酸（Asp14，D14）。利用从大肠杆菌中过表达一个合成基因获得的突变蛋白，其中将推测的与可去除铁配位的天冬氨酸残基Asp14替换为半胱氨酸，我们研究了不稳定簇的簇间转换特性。通过电子顺磁共振（EPR）和磁圆二色光谱分析表明，Asp14 --> Cys（D14C）突变体含有两个[4Fe-4S]2+/1+簇，在还原态时两者都具有S = 1/2。此外，与天然的8Fe非洲脱硫弧菌铁氧化还原蛋白III不同，发现4Fe <--> 3Fe簇间转换反应缓慢且不完全。据推测，天然蛋白中反应的可逆性是由于14位存在天冬氨酸残基，并且这个残基可能保护[3Fe-4S]簇不被进一步降解。
