Ribulose Diphosphate Carboxylase in Thiorhodaceae

Abstract

SUMMARYThe concentration of ribulose 1,5-diphosphate (RuDP) carboxylase, the enzyme which catalyses the conversion of ribulose 1,5-diphosphate + CO, to 3-phosphoglyceric acid, was partially repressed in some Thiorhodaceae organisms when these were grown on certain organic compounds. Transfer of thiosulphate-grown organisms possessing a high concentration of enzyme into growth medium containing pyruvate caused a rapid decline in carboxylase activity. In the reverse situation, pyruvate-grown organisms preferentially synthesized RuDP carboxylase when transferred to growth medium containing thiosulphate alone. The presence of thiosulphate prevented loss of carboxylase with pyruvate. The incorporation pattern of 14C02 into the ethanol-soluble compounds of organisms metabolizing thiosulphate alone was typical of autotrophic metabolism; most of the CO, was fixed via the reductive pentose cycle. The pattern of incorporation of 14C0, by organisms metabolizing pyruvate was strikingly different in that CO, entered the cell constituents predominantly via a carboxylation leading to a four-carbon product. However, even under the latter conditions, the RuDP carboxylase and the reductive pentose cycle appeared to operate to some extent, since phosphoglycerate was an early product of CO, fixation. Phosphoglycerate was an early product of 14C0, fixation by thiosulphate-grown organisms incubated with a variety of organic substrates, showing that the carboxylase and the reductive pentose cycle could function under these conditions. The addition of thiosulphate increased incorporation of 14C0, into phosphate esters by pyruvate-grown organisms incubated with pyruvate. It is concluded that ribulose diphosphate carboxylase and the reductive pentose cycle function in Thiorhodaceae even when grown on organic substrates ; its quantitative importance was not assessed. The synthesis of the carboxylase was influenced by the growth substrate.

Thiosulphate Metabolism and Rhodanese in Chromatium sp. strain D

“…The culture of Chromatium sp. strain D and its maintenance as stab cultures on the thiosulphate + salts medium have been previously described (Hurlbert & Lascelles, 1963). This organism was used throughout these studies except in the investigations with extracts where the Athiorhodaceae, Rhodospirillum rubrum and Rhodopseudomolzas spheroides were also used.…”

“…These organisms were grown anaerobically in the light on the malate + glutamate medium described by Lascelles Media. The thiosulphate + bicarbonate + salts medium used for the growth of Chromatium D was that described by Hurlbert & Lascelles (1963). In some experiments organic substrates were used in place of thiosulphate, as described previously.…”

“…The extraction procedure was repeated at least twice. The protein suspension was collected by centrifuging and the protein determination carried out as described by Hurlbert & Lascelles (1963). Protein determinations at all stages of extract fractionation after the removal of pigments from the crude extract were made by U.V.…”

“…Preparation of extracts, Organisms were normally harvested at the end of growth when largely free of endogenous sulphur by centrifuging at 2500 g for 15 min., washed twice in one-tenth the original culture volume of 0.04 M-potassium phosphate buffer (pH 6.1) and resuspended in the same buffer a t concentrations to 50 mg. protein/ml. Crude extracts were prepared by the French press or by ultrasonic treatment (Hurlbert & Lascelles, 1963). They were centrifuged at 18,000 g for 20 min.…”

Reversible inactivation and characterization of purified inactivated form I ribulose 1,5-bisphosphate carboxylase/oxygenase of Rhodobacter sphaeroides

“…These involve enzyme-mediated activation of native RubisCO by the enzyme RubisCO activase (31,40) and, in addition, reversible binding of the intracellular inhibitor 2-carboxy-arabinitol-1-phosphate to carbamylated RubisCO (3,15). In bacteria, there have been periodic indications that RubisCO might be subject to some form of modification or alteration of its activity in vivo (17,21,34), and recent work with the photosynthetic bacterium Rhodomicrobium vannielii suggests a covalent phosphorylation (26); in this latter study, however, no information relative to how phosphorylation affected RubisCO activity was presented. In other photosynthetic bacteria, particularly Rhodobacter sphaeroides and Rhodobacter capsulatus, two different forms of RubisCO are synthesized (12,13,33).…”