The prosthetic group of cytochrome a2

Abstract

A green hemin was obtained from Aerobacter aerogenes and several other bacteria, all of which contained cytochrome a2. The yield of this hemin was related to the intensity of the 630 m[mu] band of cytochrome a2. This hemin was partially purified, but its lipophilic nature and the similarity of its side chains to those of proto-hemin made complete separation from lipids or protohemin difficult. The spectroscopic properties of the hemin and of its ferrous, carbon monoxide, cyanide and pyridine compounds were examined. These are consistent with the view that this hemin is the unaltered prosthetic group of cytochrome a2. Removal of iron from hemin a2 yielded a new chlorin. This was highly purified, but still remained oily in nature. The chlorin in ether has a principal maximum of absorption at 653 m[mu], and three lesser maxima at 598, 573 and 503 m[mu]. The Soret band is at 504 m[mu]. In 8% HC1 the chlorin has its principal maximum at 630 m[mu] but this is shifted to 647 m[mu] in 16% HC1. Paper chromatography of chlorin a2 methyl ester was carried out in various solvent systems and its behavior compared with other chlorin-like pigments and with certain porphyrins. Chlorin a2 possesses at least one vinyl side chain. The presence of a formyl or carbonyl group could not be demonstrated. The chlorin was converted into a porphyrin by (a) catalytic hydrogena-tion and reoxidation, (b) the action of hydriodic acid. The principal porphyrin obtained by the action of hydriodic acid was similar to, but not identical with, monovinyl-monohydroxyethyldeuteroporphyrin. The porphyrin obtained by catalytic hydrogenation was similar to, if not identical with, mesoporphyrin. The experimental evidence is discussed, and it is concluded that chlorin a2 is a chlorin corresponding to proto-porphyrin or to a closely related vinyl-porphyrin possessing a hydroxyethyl or ethyl side chain. A molar extinction for chlorin a2 based on its similarity to certain other chlorins was assumed. From this value the molar ratio of hemin a2 to protohemin was calculated for the hemins extracted from bacteria of different cytochrome a2 and b1 content.