Proton motive force in a photodenitrifier, Rhodopseudomonas sphaeroides forma specialis denitrificans

The washed cells of Rhodopseudomonas sphaeroides f.sp. denitrificans developed a Δp of about −175 to −200 mV during denitrification in the dark and −200 to −245 mV in the light. With NO2− as the terminal acceptor, Δp was less than with NO3−, N2O, or O2. The values of Δψ in the dark were about −150 mV for NO3− and N2O and −140 mV for NO2−. During photodenitrification with NO3−, NO2−, or N2O or respiration to O2 in light, Δψ ranged between −152 and −167 mV. Like Δψ, the ΔpH was higher in light than in the dark, resulting in a 20- to 30-mV increase in Δp during illumination with NO3−, NO2−, or N2O as the acceptor. Both ΔpH and Δψ were reduced at higher pH values (≥ 7.5). Changes in pH in response to O2 in the light were less than those in the dark, indicating light inhibition of O2 respiration. The cells maintained a reasonably high Δp without addition of a substrate or when inhibitors were used; the cells retained a fairly high Δψ even in the presence of an inhibitor. However, ΔpH was appreciably lowered and in some cases it was almost abolished when either KCN, rotenone, NaN3, carbonyl cyanide m-chlorophenylhydrazone (CCCP), 2,4-dinitrophenol, N,N′-dicyclohexylcarbodiimide, antimycin A, or 2-n-heptyl-4-hydroxyquinoline-N-oxide (HOQNO) was used. The combination of an uncoupler (e.g., CCCP) and an electron transport effector (e.g., antimycin A) further reduced the ΔpH. Antimycin A and HOQNO were more effective in inhibiting photosynthetic electron transport to NO3−, NO2−, N2O, or O2 than the dark respiration to these substrates. Dibromomethylisopropyl-p-benzoquinone, a quinone antagonist, markedly reduced ΔpH in light with NO3−, NO2−, N2O, or O2 as the terminal acceptor, indicating that photosynthetically generated electrons are used for denitrification in this bacterium.