SUMMARYFermentation balances determined for different substrates in batch and continuous cultures of Lactobacillus casei revealed two pathways of pyruvate conversion by this organism, a reduction to lactate and the phosphoroclastic cleavage. Pyruvate formed anaerobically from mannitol and citrate was split by the phosphoroclastic enzyme. Lactate was the main fermentation product formed during aerobic growth on mannitol and anaerobic and aerobic growth on glucose. In glucose-limited continuous cultures pyruvate conversion was dependent on the dilution rate. At low dilution rates glucose was fermented exclusively to acetate, ethanol and formate. At high rates only small amounts of acetate, ethanol and formate were formed and lactate production was maximal. Lactate dehydrogenase of L. casei had an absolute requirement for fructose-I,6-diphosphate and manganous ions. The specific activity of lactate dehydrogenase did not differ significantly at different dilution rates. It was concluded that the intracellular level of fructose-I ,6-diphosphate controlled the pathway of pyruvate conversion. In batch cultures YATP values were between 18-2 and 20.9. No evidence for oxidative phosphorylation was found. In continuous cultures YdTP values varied from 18.7 at low dilution rates to 23'5 at high dilution rates. From the dependence of YATP on the dilution rate, a maintenance coefficient of 1-52 x I O -~ was calculated. The YATP value corrected for energy of maintenance was 24.3. The possibility that the molar growth yields were erroneously high because of assimilation of growth substrate into intracellular polysaccharides, or because of energy yield from components of the medium other than the added energy source, was excluded.

SUMMARYAmongst the family of the Enterobacteriaceae the ability to reduce tetrathionate to thiosulfate and thiosulfate to sulfite and sulfide occurs in the genera Proteus, Citrobacter and Salmonella. These reductions are coupled to a respiratory chain which functions under anaerobic conditions. Only during transport of electrons to tetrathionate oxidative phosphorylation has been demonstrated.Isolation and purification of the cytoplasmic membrane bound tetrathiohate and thiosulfate reductase from Proteus mirabilis makes clear that this bacterium forms only one enzyme for both reductions. This enzyme has a molecular weight of 133,000 daltons and can be divided into two subunits with molecular weights of 43,000 and 90,000 daltons by treatment with sodium dodecyl sulfate and 2-mercaptoethanol.The reduction of tetrathionate is activated by its primary product thiosulfate.Nitrate or oxygen represses and inactivates the tetrathionate and thiosulfate reductase. Nevertheless the smaller subunit of this enzyme appears to be formed and assembled into the cytoplasmic membranes after anaerobic growth in the presence of nitrate. EXPERIMENTAL Regulation o/the/ormation o/tetrathionate and thiosul/ate reductaseAmongst the family of the Enterobacteriaceae members of the genera Proteus, Citrobacter and Salmonella exhibit the ability to reduce tetrathionate to thiosulfate, and thiosulfate to sulfite and sulfide. The concerning specific enzyme activities for three representatives of these genera are listed in Table 1. The enzyme activities

Paracoccus denitrificans was aerobically grown in chemostat culture with succinate or gluconate as carbon source. Due to the presence of two phosphorylation sites in the respiratory chain and the absence of branching, theoretical P/O ratios of 1.71 and 1.82 were calculated for cells growing respectively with succinate and gluconate as carbon source. Using these data, 95% confidence intervals for the P/O ratio were determined, via a mathematical model, at 0.91-1.15 and 1.00-1.37 for sulphate-limited cultures, with respectively succinate and gluconate as carbon source. These results and measurements of P/O ratios in membrane particles and of proton translocation in whole cells have led to the conclusion that site I phosphorylation is affected under sulphate-limited conditions. Under conditions of carbon source-limitation the endogenous leads to H+/O ratio is about 7-8. Average values of 3.40 and 4.78 were respectively found for sulphate-limited succinate- and gluconate grown cells. For starved cells, oxidizing succinate as exogenous substrate, the leads to H+/O ratios were determined at about 3-4, independent of the growth limiting factor. It is concluded that the number of protons ejected per pair of electrons per energy-conserving site (leads to H+/site ratio) is about 3-4, instead of 2 as postulated by the chemiosmotic hypothesis.

1. Proteus mirabilis formed fumarate reductase under anaerobic growth conditions. The formation of this reductase was repressed under conditions of growth during which electron transport to oxygen or to nitrate is possible. In two of three tested chlorate-resistant mutant strains of the wild type, fumarate reductase appeared to be affected. 2. Cytoplasmic membrane suspensions isolated from anaerobically grown P. mirabilis oxidized formate and NADH with oxygen and with fumarate, too. 3. Spectral investigation of the cytoplasmic membrane preparation revealed the presence of (probably at least two types of) cytochrome b, cytochrome a1 and cytochrome d. Cytochrome b was reduced by NADH as well as by formate to approximately 80%. 4. 2-n-Heptyl-4-hydroxyquinilone-N-oxide and antimycin A inhibited oxidation of both formate and NADH by oxygen and fumarate. Both inhibitors increased the level of the formate/oxygen steady state and the formate/fumarate steady state. 5. The site of inhibition of the respiratory activity by both HQNO and antimycin A was located at the oxidation side of cytochrome b. 6. The effect of ultraviolet-irradiation of cytoplasmic membrane suspensions on oxidation/reduction phenomena suggested that the role of menaquinone is more exclusive in the formate/fumarate pathway than in the electron transport route to oxygen. 7. Finally, the conclusion has been drawn that the preferential route for electron transport from formate and from NADH to fumarate (and to oxygen) includes cytochrome b as a directly involved carrier. A hypothetical scheme for the electron transport in anaerobically grown P. mirabilis is presented.

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