Purification and characterization of cytochrome ƒ-556.5 from the blue-green alga Spirulina platensis

Abstract

The membrane-bound cytochrome ƒ-556.5 from the blue-green alga Spirulina platensis was purified to apparent homogeneity. Most of its properties are comparable to cytochrome ƒ isolated from higher plants and green algae. It is clearly distinguishable from soluble cytochrome c-554, also present in Spirulina, which probably replaces the function of plastocyanin in photosynthetic electron transport. 1. 1. The reduced form of cytochrome ƒ exhibits an asymmetrical α-band with a maximum at 556.5 nm, and a pronounced shoulder at 550 nm. The β-, γ and δ-bands coincide with those described for Scenedesmus cytochrome ƒ-553, with maxima at 524 (532), 422, 331 and a protein peak at 276 nm. The maximum of ferricytochrome ƒ is at 410.5 nm; there is no indication of a weak 695 nm band, described for soluble c-type cytochromes. The purest preparations had a δ/protein-peak ratio of 0.8; the γ/α ratio was 7.3. Formation of a pyridine hemochromogen with a maximum at 550 nm indicated a c-type cytochrome. The molar extinction coefficient at 556.5 nm is 30 200, the differential extinction coefficient 21 500. 2. 2. The molecular weight determined by gel filtration or SDS-polyacrylamide gel electrophoresis is 33 000 and 34 000, respectively. 3. 3. The redox properties differ from those described for other cytochromes ƒ isolated from green algae and higher plants: the midpoint redox potential is significantly more negative (+318 mV, pH 7.0) and from pH 6 to 10 no pH dependence is observed. 4. 4. The isoelectric point was determined at pH 3.95, which is more acidic as compared to other cytochromes ƒ. 5. 5. Comparison of the amino acid composition indicated a distant relationship to higher plant cytochrome ƒ and a closer relationship to cytochrome ƒ from green algae.