BACTEROIDS FROM SOYBEAN ROOT-NODULES - RESPIRATION AND N2-FIXATION IN FLOW-CHAMBER REACTIONS WITH OXYLEGHEMOGLOBIN

Suspensions of bacteroids prepared from root nodules of glasshouse-grown soybeans were studied in a stirred chamber supplied with variable flows of solutions containing dissolved air, oxyleghaemoglobin and various energy sources. In experiments of up to 6 h duration, several steady states were established in which frequent measurements were made of the concentration of free dissolved O2 in the range 5-200 nM and of rates of O2 consumption, CO2 efflux and N2 fixation. The principal findings were: (i) bacteroides were capable of efficient N2 fixation without the supply of exogenous energy sources when respiring at 10-50 nM free O2. This endogenous respiration was enhanced after periods of supply of succinate or malate. (ii) Exogenous energy sources were of three types, those that were poorly utilized (glucose), those that enhanced endogenous respiration and supported efficient N2 fixation (glutamate, 2-oxoglutarate) and succinate and malate, which promoted respiration but supported N2 fixation inefficiently or in some circumstances inhibited it. It is proposed that succinate and malate act primarily to increase endogenous reserves that become the principal source of reducing power for N2 fixation. (iii) Bacteroids have a terminal oxidase system with very high apparent affinity for O2 (s0.5 ~ 5-8 nM) and complex kinetics (plots of v against s are sigmoidal; n(app) > 1.8). This system was active with endogenous substrates and when glutamate or 2-oxoglutarate were supplied. With succinate or malate, respiration appeared to be the sum of endogenous activity plus O2 consumption by a system with lower affinity for O2 (K(s) = 38 nM) and simple kinetics (n(app) ~ 1). (iv) During the first hour of reactions there were changes in O2 demand and oscillations in O2 demand and CO2 efflux followed supply or withdrawal of exogenous substrates. It is proposed that these changes are examples of phenotypic plasticity in these symbiotic bacteria.