Iron-sulfur cluster-containing proteins carry out a very wide range of functions in life, ranging from electron transfer and catalysis to gene regulation and sources of sulfur.  Working Group 4 is focused on understanding the mechanisms by which iron-sulfur clusters carry out these functions. Mechanistic insight can be gained from both in vitro and in vivo approaches. In vitro work, usually with purified proteins, involves the application of a wide range of biophysical techniques, including spectroscopies (e.g. absorbance, CD, fluorescence, EPR, Mössbauer, resonance Raman, FT-IR, NRVS) structural methods (e.g. X-ray, NMR and cryo-EM), bioanalytical methods (ITC, SPR and AUC), various mass spectrometries (e.g. LC-MS, native MS), and rapid reaction kinetic methods. Studies of wild type and site-directed variants are often employed to determine residues that play important roles.  In vivo work can also take many forms, such as gene knock out, complementation and/or expression studies, proteomics, metabolomics, and two-hybrid studies. The most powerful insights are typically gained from complementary in vitro and in vivo studies.