Kurtz D M, McMillan R S, Burgess B K, Mortenson L E, Holm R H
Proc Natl Acad Sci U S A. 1979 Oct;76(10):4986-9. doi: 10.1073/pnas.76.10.4986.
The core extrusion method has been applied to the determination of the type ([2Fe-2S], [4Fe-4S]) and number of iron-sulfur centers in the FeMo proteins of the nitrogenases from Clostridium pasteurianum and Azotobacter vinelandii. The method involves extrusion with o-xylyl-alpha, alpha'-dithiol, ligand exchange of the extrusion products with p-CF3C6H4SH (RFSH), and identification and quantitation of the resultant [FenSn(SRF)4]2- complexes (n = 2,4) by 19F NMR spectroscopy. In hexamethylphosphoramide/water, 4:1 (vol/vol), 49-56% of the Fe content was extruded as [Fe4S4(SRF)4]2-, corresponding to 3,4-4.0 Fe4S4 cores per alpha 2 beta 2 subunit complex. The extruded iron does not arise from the FeMo cofactor, separate examination of which detected no extrusion products, and corresponds to 90-103% of noncofactor iron. No significant quantity of Fe2S2 cores was extruded. These results indicate the presence of four [4Fe-4S] centers per alpha 2 beta 2 subunit complex in preparations undepleted in iron. There are two main structural populations of iron atoms in these proteins, those in the cubane-type Fe4S4 cores and those in the FeMo cofactor.
核心挤压法已应用于测定巴氏梭菌和棕色固氮菌固氮酶的铁钼蛋白中铁硫中心的类型（[2Fe - 2S]，[4Fe - 4S]）和数量。该方法包括用邻二甲苯-α，α'-二硫醇进行挤压，挤压产物与对三氟甲基苯硫酚（RFSH）进行配体交换，以及通过19F核磁共振光谱法鉴定和定量所得的[FenSn(SRF)4]2-配合物（n = 2,4）。在六甲基磷酰胺/水（体积比4:1）中，49 - 56%的铁含量以[Fe4S4(SRF)4]2-形式被挤出，相当于每个α2β2亚基复合物有3.4 - 4.0个Fe4S4核心。挤出的铁并非来自铁钼辅因子，单独检测铁钼辅因子未发现挤出产物，且挤出的铁相当于非辅因子铁的90 - 103%。未挤出大量的Fe2S2核心。这些结果表明，在铁未耗尽的制剂中，每个α2β2亚基复合物存在四个[4Fe - 4S]中心。这些蛋白质中的铁原子主要有两种结构群体，即立方烷型Fe4S4核心中的铁原子和铁钼辅因子中的铁原子。
