Identification of the CO-binding cluster in nitrogenase MoFe protein by ENDOR of Fe-57 isotopomers
Identification of the CO-binding cluster in nitrogenase MoFe protein by ENDOR of Fe-57 isotopomers
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DOI：
10.1021/ja961289g
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发表时间：
1996-09-11
影响因子：
15
通讯作者：
Hoffman, BM
中科院分区：
文献类型：
--
作者：
Christie, PD;Lee, HI;Hoffman, BM
氮酶MoFe蛋白的x射线结构确定了其多金属辅助因子p -簇（Fe8S7-8）和femo辅助因子（MoFe7S9：高柠檬酸盐）的组织和结构。然而，直到最近，还不可能检测或表征与功能酶相互作用的底物或抑制剂。在本研究中，我们使用Fe-57 ENDOR研究了一组新的Fe-56、Fe-57 MoFe蛋白同位素体的氧化石墨烯抑制转换状态，包括那些这两个簇被选择性和均匀标记的。go抑制MoFe蛋白在低CO压和高CO压下表现出两种不同的EPR信号。Fe-57的测量，以及早期的C-13 ENDOR对束缚(CO)-C-13的研究（波洛克，r.c.；李，h.i.；卡梅隆，l.m.；德罗斯，v.j.；黑尔斯，b.j.；奥姆-约翰逊，w.h.；霍夫曼，b.m.j.）。陈省身。Sec. 1995, 117, 8686-8687)的研究表明，这两个EPR信号是由go结合的femo - CO因子产生的，在一种情况下有一个结合的CO，而在另一种情况下有两个结合的CO，它们进一步提供了对抑制物结合簇的性质的初步见解。
The X-ray structure of the nitrogenase MoFe protein has established the organization and architecture of its multimetallic cofactors, the P-cluster (Fe8S7-8) and the FeMo-cofactor (MoFe7S9:homocitrate). Nonetheless, until recently it has not been possible to detect or characterize a substrate or inhibitor interacting with the functioning enzyme. In the present study we have used Fe-57 ENDOR to study the GO-inhibited turnover states of a novel suite of Fe-56,Fe-57 isotopomers of the MoFe protein, including those in which these two clusters are selectively, as well as uniformly, labeled. GO-inhibited MoFe protein exhibits two distinct EPR signals, one under low and another under high CO pressure. The Fe-57 measurements, along with an earlier C-13 ENDOR study of bound (CO)-C-13 (Pollock, R. C.; Lee, H. I.; Cameron, L. M.; DeRose, V. J.; Hales, B. J.; Orme-Johnson, W. H.; Hoffman, B. M. J. Am. Chern. Sec. 1995, 117, 8686-8687), show that the two EPR signals arise from GO-bound FeMo-cofactor, in one case with one bound CO and in the other with two bound CO, and they further provide initial insights into the properties of the inhibitor-bound cluster.