Functional similarities of nitrate reductase from yellow lupine bacteroids to bacterial denitrification systems

Abstract

The nitrate-reducing activity of root nodules of field-grown yellow lupine plants (Lupinus luteuscv. Ventus) was found to be about 25 and 100 times higher than that of roots and leaves, respectively. At least 97 percnt; of nodule nitrate reductase (NR) activity proved to be of bacteroidal origin. Data from the present work indicates that nitrate reductase (EC 1. 7. 99. 4) of symbiotic Bradyrhizobium sp. (Lupinus) cells possess some functional similarities to bacterial denitrification systems. The enzyme occurs in association with membranes, and electron transport from NADH to nitrate is dependent on the mediation of membrane components. Another feature in common with bacterial nitrate dissimilation is inhibition of bacteroidal nitrate reductase activity under aerobic conditions. Membrane permeabilization significantly alleviated this inhibitory effect. It was found that enzyme activity is not nitrate-dependent and peaks at pH 5.5-6.5. Rotenone inhibition indicated that NADH-quinol oxidoreductase, not succinate dehydrogenase, is the major participant in a very efficient succinate-dependent nitrate reductase activity. Pyruvate and malate were also highly effective in nitrate reduction. By contrast, oxaloacetate and 2-oxoglutarate were not efficient, even in combination with pyruvate, which suggests that this effect could not be caused by acetyl-SCoA limitation. These results suggest that a large part of the TCA cycle may not be necessary for nitrate reduction in yellow lupine bacteroids. The possible alternative pathways, which could provide electrons from pyruvate and malate to nitrate reductase, are discussed.