Achieving\nStable Partial Denitrification by Selective\nInhibition of Nitrite Reductase with the Biosafe Aprotic Solvent DMSO

The recently proposed partial denitrification\n(PD), terminating\nnitrate reduction to nitrite, has been regarded as a promising alternative\nto nitrite supplying for anammox bacteria. The most important aspect\nof the PD process for engineering application is the stable and continuous\nsupply of nitrite. However, the activity of nitrate reductase is often\nhigher than that of nitrite reductase (NIR), making it difficult to\naccumulate nitrite during the denitrification process. Herein, a strategy\nfor achieving efficient and stable partial denitrification using the\nbiosafe additive dimethyl sulfoxide (DMSO) was constructed, and the\nmechanism of DMSO inhibiting NIR was analyzed. DMSO addition reduced\nthe expression of <i>NIR</i> gene, and 1% DMSO addition\ncan significantly inhibit NIR enzyme activity to achieve a stable\nPD process. When the DMSO concentration increased to 3.5%, the NIR\nenzyme activity was almost inhibited with the enzyme activity of only\n0.95 mg nitrite/min. However, the addition of DMSO has almost no inhibitory\neffect on the nitrate reductase (NAR) enzyme. The affinity constant\nof DMSO with the NAR enzyme is −2.4 kcal/mol, while the affinity\nconstant of DMSO with the NIR enzyme is as high as −3.1 kcal/mol.\nDMSO shows a higher affinity for NIR. Moreover, DMSO and nitrite occupy\nthe same catalytic cavity in the NIR enzyme, which is the fundamental\nreason why DMSO selectively inhibits the NIR enzyme. This study provides\na new idea for realizing efficient and stable partial denitrification\nfunction.