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SLIDE SHOW III Rescuing E325D

SLIDE SHOW III Rescuing E325D. A. Mutant E325D is partially uncoupled and has lower K D for substrate. +. -. -. -. IX. R 302. D 325. X. H 322. VIII. K 319. H + o. E 269. +. D 240. R. +. C. 144. VII. 148. -. V. E 126. IV. +. -. -. -. -. +. +.

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SLIDE SHOW III Rescuing E325D

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  1. SLIDE SHOW III Rescuing E325D

  2. A. Mutant E325D is partially uncoupled and has lower KD for substrate. + - - - IX R302 D325 X H322 VIII K319 H+o E269 + D240 R + C 144 VII 148 - V E126 IV

  3. + - - - - + + B. Substrate binds from the outside with lower affinity. IX R302 D325 X H322 VIII K319 H+o E269 D240 R C 144 VII So 148 V E126 IV

  4. C. Binding of substrate causes a conformational change disrupting the interaction of Glu269 and His322, …. + - - - - + IX R302 D325 X H322 VIII K319 H+o E269 + D240 R C 144 VII So 148 V E126 IV

  5. D. ... causes substrate to become accessible to the internal surface, and the proton moves to His322/Asp325. + - - - IX R302 H+ D325 VIII H322 X K319 E269 + D240 R + C 144 Si VII 148 - V E126 IV

  6. E. Rotation of Helix X continues towards the hydrophobic phase of the membrane, + - - - IX R302 H+ D325 VIII H322 X K319 E269 + D240 R + C 144 Si VII 148 - V E126 IV

  7. + - - - F. raising the pKa of Asp325. H+ IX R302 VIII D325 H322 K319 X E269 + D240 R + C 144 Si VII 148 - V E126 IV

  8. - - - G. Substrate dissociates. H+ IX R302 + VIII D325 H322 K319 X E269 + - Si D240 R + C Si Si 144 VII 148 V E126 IV

  9. H. By introducing a Gly or Ala residue at the periplasmic end of helix X, Asp325 re-juxtaposes with Arg302 without improving affinity for substrate. + - - - H+ IX R302 D325 VIII H322 K319 X E269 + V316G/A D240 R + C 144 VII 148 - V E126 IV

  10. I. The proton is released to the inside with increased efficiency, ….. + - - - - + + IX R302 D325 X H322 H+i VIII K319 V316G/A E269 D240 R C 144 VII 148 V E126 IV

  11. J. and the permease relaxes into the ground state and becomes protonated from the outside. + - - - + IX R302 D325 X H322 VIII K319 H+o V316G/A E269 D240 R + C 144 VII 148 - V E126 IV

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