Molecular evolution of a membrane protein: CLC channel from/to CLC ant…

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  • 2017-01-06


[BK21 Plus Seminar]
                    ▶Subject: Molecular evolution of a membrane protein: CLC channel from/to CLC antiporter
                    ▶Speaker: Hyun-Ho Lim, Ph.D. (Korea Brain Research Institute (KBRI))

                    ▶Date: 4:00PM/Oct. 18(Tue.)/2016
                    ▶Place: Life Science Bldg. #104
                    Ion channels and ion transporters are essentially membrane-embedded enzymes, which catalyze the translocation of substrates (ions) in and out of cell membrane. However, these two proteins are operating with very different biophysical principles. In contrast to ion channels allow electrochemically downhill movement of ions upon stimulation, membrane transporters requires free energy input to transport ion in energetically uphill direction. Thus, it had been generally believed that these two are structurally and evolutionarily unrelated protein superfamily, until Chris Miller and his colleagues revealed that CLC family proteins turned out to be either Cl- channel or Cl-/H+ antiporter.
 The CLC superfamily can be found in virtually all organisms from bacteria to human. In mammal, genetic mutations in the CLC genes are linked to the various diseases such as myotonia, deafness, epilepsy, leukodystrophy, kidney malfunctions and lysosomal storage disease. CLC proteins also play key roles in the physiology of other organisms: nitrate uptake for nitrogen fixation in plants, gastric acid-resistance in enteric bacteria and fluoride (F-)-resistance mechanisms of unicellular microorganisms such as pathogenic bacteria and eukaryotic parasites.
 In E.coli, two CLC proteins, CLC-ec1 and CLC-ec2 are both functionally important for bacteria to survive extreme acid challenge: the double knockout cannot support survive at pH 2.5, but the presence of either CLC gene equally the double knockout mutant. However, in vitro purified and reconstituted CLC-ec2 shows much lower transport activity than CLC-ec1. How can CLC-ec2 have such a physiological contribution with such a functional difference?
 We have been collecting a line of evidences which suggests a likely answer: CLC-ec2 is activated below pH 3, where CLC-ec1 begins to shut down and CLC-ec2 activity is stimulated upon voltage changes to which CLC-ec1 does not respond. Moreover, CLC-ec2 swaps one proton with ~10 Cl- in the reconstituted proteoliposome, but the stoichiometry can be interestingly reduced to 1 to 4 (H+/Cl-) by transmembrane voltage stimulus and acidification. What can change the coupling ratio of transported ions in CLC-ec2? Systematic mutagenic approaches produce results that voltage-dependent H+ translocation could be the culprit of changing coupling ratio from. In order to obtain structural and mechanistic insights, x-ray crystallographic work propel the structural determination of CLC-ec2 protein. These results imply that the voltage-dependent H+ transport and the presence of Clslippage state might be the molecular determinants to build ‘ion channel’- and ‘transporter’-type CLC proteins.

                ▶Inquiry: Prof. Cheol-Sang Hwang (279-2352)
                      * This seminar will be given in English.
                  please refrain from taking photos during seminars. *