Abstract
(57)Fe Mӧssbauer spectroscopy is unparalleled in the study of Fe-S cluster-containing proteins because of its unique ability to detect all forms of iron. Enrichment of biological samples with the (57)Fe isotope and manipulation of experimental parameters such as temperature and magnetic field allow for elucidation of the number of Fe-S clusters present in a given protein, their nuclearity, oxidation state, geometry, and ligation environment, as well as any transient states relevant to enzyme chemistry. This chapter is arranged in five sections to help navigate an experimentalist to utilize (57)Fe Mӧssbauer spectroscopy for delineating the role and structure of biological Fe-S clusters. The first section lays out the tools and technical considerations for the preparation of (57)Fe-labeled samples. The choice of experimental parameters and their effects on the Mӧssbauer spectra are presented in the following two sections. The last two sections provide a theoretical and practical guide on spectral acquisition and analysis relevant to Fe-S centers.