The interaction of nitric oxide (NO) with mitochondria is of pathological significance and is also a potential mechanism for the regulation of mitochondrial function. Some of the ways in which NO may affect mitochondria are by reacting with low-molecular-weight thiols such as glutathione and with protein thiols. However, the detailed mechanisms and the consequences of these interactions for mitochondria are uncertain. Here we review mitochondrial thiol metabolism, outline how NO and its metabolites interact with thiols, and discuss the implications of these reactions for mitochondrial and cell function.