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Bacteroids from soybean root nodules: accumulation of poly-β-hydroxybutyrate during supply of malate and succinate in relation to N2 fixation in flow-chamber reactions

A liquid reaction medium containing dissolved air and oxyleghaemoglobin was supplied to bacteroids confined in a stirred reaction chamber with no gas phase. The relative oxygenation of leghaemoglobin, dissolved CO2 and NH3 from N2 fixation were monitored before, during (when N2 fixation was diminished) and after the supply of [14C]malate and succinate. The amounts and location of 14C-labelled products were determined in bacteroids recovered from the chamber and 14CO2 was measured in the effluent. During the supply of [14C]succinate, the bacteroids accumulated most 14C from C2 and C3 whereas 14CO2 arose primarily from C1 and C4. This suggested that production of pyruvate from malate via malic enzyme was a central feature. For up to 80 min after removal of [14C]malate and succinate, CO2 arose from stored carbon and N2 fixation was enhanced; during this period 14C continued to accumulate in poly-β-hydroxybutyrate, until this accounted for about 90% of bacteroid 14C. Thereafter CO2 was evolved with steady, low radioactivity and enhanced N2 fixation continued. These results supported the proposal that poly-β-hydroxybutyrate, which is present in large amounts (50-70% of dry mass) in these bacteroids, is a mobilizable, energy-yielding reserve that provides endogenous substrates for support of N2 fixation when exogenous substrates are not available.