Integral caa₃-Cytochrome c Oxidase from Thermus thermophilus: Purification and Crystallization

Cytochrome c oxidase is a respiratory enzyme catalyzing the energy-conserving reduction of molecular oxygen to water—a fundamental biological process of cell respiration. The first crystal structures of the type A cytochrome c oxidases, bovine heart and Paracoccus denitrificans cytochrome c oxidases, were published in 1995 and contributed immensely to the understanding of the enzyme’s mechanism of action. The senior author’s research focus was directed toward understanding the structure and function of the type B cytochrome c oxidases, ba₃-oxidase and type A2 caa₃-oxidase, both from the extreme thermophilic bacterium Thermus thermophilus. While the ba₃-oxidase structure was published in 2000 and functional characterization is well-documented in the literature, we recently successfully solved the structure of the caa₃-nature made enzyme-substrate complex. This chapter is dedicated to the purification and crystallization process of caa₃-cytochrome c oxidase.