The Purification and Properties of Cytochrome o from Vitreoscilla

Abstract

Abstract By freezing and thawing a suspension of Vitreoscilla cells in the presence of 1% sodium deoxycholate, followed by a protamine sulfate step, an ammonium sulfate step, Sephadex G-100 chromatography, and TEAE-cellulose chromatography, two CO-binding pigments have been purified. One (Fraction I) has two α bands and two Soret bands in liquid nitrogen difference spectra and thus appears to possess two heme groups. A molecular weight of 27,000 was determined by chromatography on Sephadex G-75. Protoheme was shown to be the prosthetic group of this pigment, and the Eo' was found to be +0.10 volt. It is only very slowly autoxidizable. The other pigment (Fraction II) is very autoxidizable and has an Eo' of -0.09 volt. The prosthetic group of this protein is also protoheme, and it has a molecular weight of 22,500. The properties of both pigments are discussed in relation to their possible roles as the terminal oxidase.