Dicarboxylic AcidTransport inMembraneVesicles from Bacillus subtilis

electron donorsystemascorbate-phenazine methosulfate. Thedicarboxylic acids areaccumulated inunmodified form. Inhib- itors oftherespiratory chain, sulfhydryl reagents, anduncoupling agents in- hibit theaccumulation ofthedicarboxylic acids. Theaffinity constants for transport ofL-malate, fumarate, andsuccinate are 13.5, 7.5,and4.3AM, respectively; these values areseveralfold lowerthanthose reported previously forwholecells. Active transport ofthese dicarboxylic acids occursviaonehighly specific transport system asisindicated bythefollowing observations. (i)Each dicarboxylic acidinhibits thetransport oftheothertwodicarboxylic acids competitively. (ii) Theaffinity constants determined fortheinhibitory action are verysimilar tothose determined forthetransport process.(iii) Eachdicarboxylic acidexchanges rapidly witha previously accumulated dicarboxylic acid. (iv) Othermetabolically andstructurally related compounds donotinhibit transport ofthese dicarboxylic acids significantly, except forL-aspartate andL-glutamate. However, transport ofthesedicarboxylic aminoacids ismediated byan inde- pendent systembecause membranevesicles fromB.subtilis 60346, lacking functional dicarboxylic aminoacid transport activity, accumulate theC4dicar- boxylic acids ateven higher ratesthanvesicles fromB.subtilis W 23.(v)A constant ratio exists between theinitial ratesoftransport ofL-malate, fumarate, andsuccinate inallmembranevesicle preparations isolated fromcells grown on various media. Thishigh-affinity dicarboxylic acid transport system seems tobe present constitutively inB.subtilis W23. Active transport ofmetabolites across thecy- toplasmic membrane hasreceived considerable attention inthelastfewyears. Moststudies havebeenconcerned withtransport systems for aminoacids andcarbohydrates, andonlyre- cently hasinterest beendirected towards trans- portsystems fortricarboxylic acidcycle inter- mediates. Several ofthesetricarboxylic acid cycle intermediates appear tobeaccumulated viaspecific andinducible transport systems. Studies inwholecells, forinstance, demon- strated thepresence ofspecific andinducible uptake systems forcitrate inKlebsiella aero- genes (4, 28), Pseudomonas spp.(20), andBacil- lussubtilis (32). InSalmonella typhimurium there appear tobethreespecific andinducible