Effect of Light on the Tricarboxylic Acid Cycle in Scenedesmus

Abstract

An attempt was made to determine the effect of light on the operation of the tricarboxylic acid cycle in a green alga. Scenedsmus obliquus was incubated in the dark or under photosynthetic conditions with acetate-1- or -2-C14 and pyruvate-3-C14. The specific activities and distribution of C14 within citrate, [alpha]-ketoglutarate, succinate, malate, and glutamate were determined. The findings established that S. obliquus possessed a classical citric-acid cycle. The results also indicated that light had no detectable effect on the turnover of the cycle. Acetate apparently entered the cycle exclusively as acetyl-CoA. Isotope from acetate-2-C14 was concentrated in the appropriate C atoms of citrate, a-ketoglutarate, succinate, malate, and glutamate as predicted by the classical Krebs cycle and spread from the noncarboxyl to the carboxyl C atoms at equivalent rates under both light and dark conditions. Isotope from acetate-1-C14 was found exclusively in the carboxyl carbons and isotopic equilibrium of the cycle acids was approached within 3 minutes in both the light and dark. The specific activity of the CO2 evolved in the dark from acetate-1-C14 was 30% of that of the tertiary carboxyl C of citrate and the [alpha]-carboxyl C of [alpha]-ketoglutarate. Pyruvate entered the cycle as acetyl-CoA and also by a pathway involving a carboxylation reaction, the latter pathway being markedly enhanced by light. The malic acid formed in the light by the carboxylation reaction was not in euqilibrium with other intermediates of the tricarboxylic acid cycle, although the intramolecular distribution data indicated that it did enter the cycle. Glutamic acid approached rapid isotopic equilibrium with [alpha]-ketoglutarate in the light and dark despite the fact that the concentration of glutamic acid was 10-20-fold greater than [alpha]-ketoglutarate. Most of the glutamic acid was derived from the tricarboxylic acid cycle.