Cholera is an acute, diarrheal disease caused by infection of the intestine with the gram-negative, motile bacterium Vibrio cholerae. After colonizing the small intestine, V. cholerae secretes cholera toxin (CTX), a protein that ribosylates the Gsa subunit of a G protein, thereby inhibiting its GTPase activity. This causes the dysregulation of a cellular chloride ion channel, resulting in efflux of ions and water from the infected enterocytes into the intestinal lumen. Studies have implicated sodium bioenergetics in the pathogenesis of V. cholerae in humans. An electrochemical gradient of Na+ known as the "sodium motive force" provides V. cholerae with the energy for key functions. The gradient is predominantly established by the Nat-pumping NADH:quinone oxidoreductase (Na+-NQR) enzyme. Na*-NQR is a multi-substrate enzyme comprised of six distinct subunits designated A-F (Table 1) that are encoded within the same operon. In a process similar to proton pumping by Complex I of the electron transport chain, Na+-NQR couples the movement of electrons from an NADH donor with ion transport. Table 1 Biochemical Features of V. cholerae Na+-NQR Subunits Number of amino Molecular acid weight Property residues (kDa) NgrA NqrB NqrC NqrD NqrE NgrF 426 415 257 210 198 408 1/Vo (1/μM/min) 1/[NADH] pl 47.6 6.30 47.4 7.16 28.6 7.30 22.8 8.50 20.5 5.35 47.1 5.25 Researchers investigated the kinetic features of Nat-NQR. Figure 1 shows a double reciprocal plot of 1/V, versus 1/concentration of one substrate, NADH, at three concentrations (1X, 2X, 3X) of the second substrate, quinone. Reactions were performed in conditions where NaCl was saturating. [quinone] Number of transmembrane helices Figure 1 Double reciprocal plot of Na*-NQR activity at three concentrations (1X, 2X, 3X) of quinone O A. Decreased activity of Na*-NQR O B. Decreased production of ATP O C. Decreased pH of the periplasm O D. Decreased consumption of 0₂ O A. NqrA and NqrB O B. NqrC and NqrE O C. NqrD and NqrF O D. NqrE and NqrF 0 8 2 5 6 1 What is the most likely effect of adding a sodium ionophore to a culture of V. cholerae? Which two subunits of Nat-NQR can be separated by gel filtration but NOT by ion exchange chromatography? O A. Malate dehydrogenase OB. Succinate dehydrogenase O C. Isocitrate dehydrogenase O D. A-Ketoglutarate dehydrogenase Which enzyme of the citric acid cycle is NOT directly involved in generation of the dinucleotide required for Nat-NQR activity? Based on information presented in Table 1, which relationship between pH and charged functional groups is accurate? O A. At pH 8.50, the ratio of the cationic to anionic functional groups in the NqrD subunit is equal to 1. O B. At pH 7.00, half of the functional groups in the NqrE subunit are protonated. O C. At pH 6.30, the net charge of the cationic and anionic functional groups in the NqrA subunit is equal to 1. O D. At pH 6.00, the majority of the functional groups in the NgrF subunit are protonated.