Ligand binding and substitution reactions are important for metalloprotein folding and function. The heme sensor of a methyl-accepting chemotaxis GSU0935 is a c-type cytochrome from the bacterium Geobacter sulfurreducens. The heme domain switches one of its axial ligands from H(2)O to a low-spin ligand, presumably Met, upon reduction. The study analyzes the stability and folding kinetics of the ferric domain. Guanidine hydrochloride denaturation yields the low-spin heme species arising from coordination of the ferric heme by non-native His residues. The population of the low-spin species further increases and then declines during protein refolding. Kinetics and mutational effects suggest that His54, from the N-terminal region of the domain, is the transient ligand to the heme. The capture and release of a non-native ligand within the compact partially-folded structures illustrates the flexibility of the heme environment in GSU0935, which may relate to the domain sensor function.