IDENTIFICATION OF THE CO-BINDING CLUSTER IN NITROGENASE MOFE PROTEIN BY ENDOR OF FE-57 ISOTOPOMERS

The X-ray structure of the nitrogenase MoFe protein has established th e organization and architecture of its multimetallic cofactors, the P- cluster (Fe8S7-8) and the FeMo-cofactor (MoFe7S9:homocitrate). Nonethe less, until recently it has not been possible to detect or characteriz e 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 MoF e protein, including those in which these two clusters are selectively , as well as uniformly, labeled. GO-inhibited MoFe protein exhibits tw o distinct EPR signals, one under low and another under high CO pressu re. 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 G O-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.