Active Transport by Membrane Vesicles from Anaerobically Grown Escherichia coli Energized by Electron Transfer to Ferricyanide and Chlorate

Abstract

Active transport of amino acids by membrane vesicles from Eschrricizicr cnli, grown anaerobically on glucose in the presence of nitrate, can be energized under anaerobic conditions by electron transfer in the nitrate respiration system with formate as electron donor and nitrate as acceptor. A high rate of amino acid transport is also obtained under anaerobic conditions by electron transfer from formate to the nitrate analogue chlorate or to the membrane-impermeable electron acceptor ferricyanide. Electron transfer from formate to nitrate results in the generation of an electrical potential as is indicated by the uptake of the lipophilic cation triphenylmethylphosphonium.Ferricyanide accepts electrons from at least two sites of the nitrate respiration system. One of these sites appears to be nitrate reductase, because cytochrome 6, reduced by formate, is completely reoxidized by ferricyanide and glutamate transport energized by formate plus ferricyanide and formate plus nitrate are affected by the same electron transfer inhibitors. A second site of electron transfer to ferricyanide appears to be located prior to nitrate reductase in the nitrate respiration system, since formate is oxidized at a higher rate in the presence of ferricyanide than with nitrate while formate/ferricyanide energizes transport of amino acids at a lower rate than formate/nitrate. Moreover, electron transfer inhibitors block electron transfer from formate to nitrate to a significantly higher extent than from formate to ferricyanide. The effects of irradiation of the membrane vesicles with near ultra-violet light suggest that quinones play an essential role in the electron transfer from formate to nitrate or ferricyanide. Irradiation blocks completely formate-dependent nitrate and ferricyanide reduction and active transport driven by formate/nitrate and formate/ferricyanide, but has hardly any effect on the activity of formate dehydrogenase and on ascorbate/phenazine methosulphatejoxygen-driven transport. Similar effects of ferricyanide have been observed in membrane vesicles from E. coli, grown anaerobically in the presence of fumarate. In these membrane vesicles a high rate of lactose and triphenylmethylphosphonium uptake under anaerobic conditions is obtained by electron transfer from glycerol 1 -phosphate to fumarate and also to ferricyanide and evidence has been presented for the involvement of cytochromes in these electron transfers.Active transport of many solutes by cytoplasmic membrane vesicles from a variety of bacteria can be energized by electron transfer in the absence of ATP or other nucleoside triphosphates [l -61. In membrane vesicles from Escherichia coli three distinct electron transfer systems can supply the energy for active transport of solutes. Active transport is coupled to the oxidation of D-lactate or phenazine methosulphate via the respiratory chain to oxygen as terminal electron acceptor in membrane vesicles from aerobically grown cells [l-41. Growth of E. coli under anaerobic conditions in the presence of nitrate ...

Dissimilatory Fe(III) and Mn(IV) reduction.

“…Ferricyanide can accept electrons from electron transport chains in membrane vessicles of B. subtilis (32) and E. coli (39). It is possible that the Fe(III)-quinate that was used in the proton translocation studies can interact with electron transport components in a similar manner.…”

Generation of an Electrochemical Proton Gradient by Nitrate Respiration in Membrane Vesicles from Anaerobically Grown Escherichia coli

“…Membrane vesicles prepared from anaerobically grown E. coli concentrate L-glutamate 230-fold under anaerobic conditions in the presence of formate as electron donor and nitrate as acceptor (Fig.4A) [22] and 500-fold under aerobic conditions with axorbate/ phenazine methosulphate (Fig. 4B).…”

“…It has been shown previously that electron transfer in the nitrate respiration system supplies the energy for active t I'LL nsport of amino acids under anaerobic conditions [7.17-191. Experiments performed with spheroplasts by Garland et al [20, 211 and studies in membrane vesicles [22] with the membrane-impermeable electron acceptor ferricyanide suggested that components of the nitrate respiration system are exposed to different sides of the membrane. These observations and the demonstration that nitrate respiration results in the translocation of protons across the membrane [20] and in the generation of a A$ [7,22] indicated that an electrochemical proton gradient might be the driving force for active transport under anaerobic conditions.…”

Dissimilatory Fe(III) and Mn(IV) Reduction

“…Ferricyanide can accept electrons from electron transport chains in membrane vessicles of B. subtilis (32) and E. coli (39). It (11).…”