How Hemorrhagic E. coli Resists the Acid Environment orihuea 300- of the Stomach pH Sensitivity of the Glu-GABA Antiporter GadC AdiC 250 20tdmeM er Recent years have been marked by a series of food poisoning outbreaks involving hemorrhagic (producing internal bleeding) strains of the bacterium Escherichia coli (E. coli). Bacteria are often a source of food poisoning. 9 91A 200 blulA e1A onsdmmaM lasigoioti esle 242 typically milder infections caused by food-borne strepto- coccal bacteria. Less able to bear the extremely acidic conditions encountered by food in the human stomach (pH 2), E. coli has not been as common a problem. The hemorrhagic strains of E. coli responsible for recent out- breaks seem to have evolved more elaborate acid-resistancen6 150 bigiodiy ot a prt b Esbom 100 os il bnieis 19 ainD o 50 dmatstull o Sshoqmohetat mott beoidmeateA e u A0B 6 wimomany 97lid bigilodgaordq 8 7 opm 5 08.200000 g0nsi or 79m usku I15obos.Tw totuk systems. pH n pcg How do hemorrhagic E. coli bacteria survive in the ww.sinowstinloteuigs i acid environ ment of the stomach? The problem they face, ule to in essence, is that they are submerged in a sea of hydrogen ordt dhiw bateibozzn 96 rfsirdwanistonn ansndmom len no abiqiloovlg bns anisionqo .c ncidmsM onltrem ohamvorobi lloo en 1o 1sdi so ions, many of which diffuse into their cells. To rid them-n Analysis dr selves of these excess hydrogen ions, the E. coli cells use amom clever system to pump hydrogen ions back out of their E.A.Z T93 1. Applying Concepts ivo19 abiqiloriqeori9 S.e a. Variable. In the graph, what is the dependent o l61uUe variable? abiga noz el cells. 9TU . anT 20 meM First, the hemorrhagic E. coli cells take up cellular m5b no hydrogen ions by using the enzyme glutamic acid decarbox- 10 b. Substrate. What is a substrate? In thisslia biqil eriT CEa ow1 lo bozoqo.ou iaueensta ylase (GAD) to convert the amino acid glutamate to investigation, what are the substrates that are nodusd y-aminobutyric acid (GABA), a decarboxylation reaction that 13OUDE0 05e nomulo2 pinotoer accumulatings 1sloqnon io v odipilirdqoiluprm pH. What is the difference in hydrogen VEWB 109IO V bn ion concentration between pH 5 and pH o TOITSini pgnon How many times more (or less) is that? Explain.w b a lo gnibnod nogoibvSos consumes a hydrogen ion. lens1169.FU59 Uoloz Second, the hemorrhagic E. coli export this GABA C. from their cell cytoplasm using a Glu-GABA antiporter called GadC (this transmembrane protein channel is called an antiporter because it transports two molecules across the 2. Interpreting Data m arli 2q91 1 a. Does the amount of amino acid transported ogiino n in the 10-minute experimental interval s201ibn uso (expressed as substrate accumulation) vary no with pH for the arginine-transporting AdiC bosr antiporter? For the glutamate-transportingls Su GadC antiporter? b. Compare the amount of substrate accumulated by AdiC in 10 minutes at pH 9.0 with that accumulated at pH 5.0. What fraction of the vitA 6T membrane in opposite directions). However, to survive elsewhere in the human body, it ph is important that the Glu-GABA antiporter of hemorrhagica.e E. coli not function, lest it short-circuit metabolism. To see if the GadC antiporter indeed functions only in acid environ- ments, investigators compared its activity at a variety of pHsbeno nib to with that of a different amino acid antiporter called AdiC, which transports arginine out of cells under a broad range of conditions. The results of monitoring transport for TA 20/u tS1stnl to onA low pH activity is observed at the higher dmsM CE8 pH? 9uun 10 minutes are presented in the graph. c. In a similar fashion, compare the amount or ) substrate accumulated by GadC at pH 9.O with that accumulated at pH 5.0. What fraction of the low pH activity is observed at the higher pH? Making Inferences Would you say that the GadC antiporter exhibits the same pH dependence as the AdiC antiporter? If not, which antiporter is less active at nonacid pHs? 4. Drawing Conclusions Is the glutamate-GABA antiporter GadC active at nonacid pHs? toM ot afnotbaio nola9aU hoqans bslquoo .e.e anolber noitettneono ieriT JeriepA 2pluoelo boasslen creng srlnorw auo9o nogz0 J0Oonos Outside S OURS nogen of b0 ategib.snside noiba ul aiaieng 0-0 7no cell cell toonib ati200 3. uoslorm (1 2onidmaMo10 alghetoly 1 DMo anst Oktue abloA GABA ses.tiiW nioto pe/ vnko JED 2on on 90 ris aiaotyb ta.e aeobns nl 5. Further Analysis The GadC antiporter also em lisb tobieoy dot or lo Glutamate VD u ww.askanolo transports the amino acid glutamine (Gln). Do you think this activity has any role to play in combating low pH environments? How would you test this hypothesis? wworg uebetoloiesvdenam Chapter 5 Membranes 111 of power th WH K Substrate accumulation (nmol per mg protein) nquiry & Analysis