Comparative Studies on the Properties of Two Ferredoxins fromPisum sativumL.

Abstract

Two ferredoxins in approximately equal amounts were isolated from 3 week old Pisum sativum L. seedlings. Both ferredoxins had identical absorption spectra with maxima at 276, 327, 424, and 468 nm in the oxidized state, and each possessed a single 2Fe-2S active centre. The isoelectric points of the two ferredoxins were both at pH 3·3, and mixtures could not be separated by isoelectric focusing on polyacrylamide gels. The midpoint redox potentials of the ferredoxins were close to −415 mV, but they differed slightly in their biological activity. Ferredoxin I was slightly the more active of the two in catalysing NADP⁺ photoreduction by Pisum or Hordeum chloroplasts whereas ferredoxin II was more active in catalysing the oxidative cleavage of pyruvate by extracts of Clostridium pasteurianum. Though the molecular weights of the ferredoxins determined by ultracentrifugation were the same within experimental error, the amino acid compositions showed marked differences. The N-terminal 40 amino acid residues of ferredoxins I and II were determined by means of an automatic sequencer. There were 15 differences, suggesting that gene duplication had occurred early in evolutionary time. Ferredoxin I appears to be more closely related to the other angiosperm ferredoxins since it differed in only 6 positions compared with the corresponding sequence for Medicago sativa (alfalfa) ferredoxin. The ratio of the two ferredoxins in Pisum sativum was shown to be dependent on the age of the seedlings and environmental growth conditions.