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2bs3
From Proteopedia
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==Overview== | ==Overview== | ||
| - | Reconciliation of apparently contradictory experimental results obtained | + | Reconciliation of apparently contradictory experimental results obtained on the quinol:fumarate reductase, a diheme-containing respiratory membrane protein complex from Wolinella succinogenes, was previously obtained by the proposal of the so-called "E pathway hypothesis." According to this hypothesis, transmembrane electron transfer via the heme groups is strictly coupled to cotransfer of protons via a transiently established pathway thought to contain the side chain of residue Glu-C180 as the most prominent component. Here we demonstrate that, after replacement of Glu-C180 with Gln or Ile by site-directed mutagenesis, the resulting mutants are unable to grow on fumarate, and the membrane-bound variant enzymes lack quinol oxidation activity. Upon solubilization, however, the purified enzymes display approximately 1/10 of the specific quinol oxidation activity of the wild-type enzyme and unchanged quinol Michaelis constants, K(m). The refined x-ray crystal structures at 2.19 A and 2.76 A resolution, respectively, rule out major structural changes to account for these experimental observations. Changes in the oxidation-reduction heme midpoint potential allow the conclusion that deprotonation of Glu-C180 in the wild-type enzyme facilitates the reoxidation of the reduced high-potential heme. Comparison of solvent isotope effects indicates that a rate-limiting proton transfer step in the wild-type enzyme is lost in the Glu-C180 --> Gln variant. The results provide experimental evidence for the validity of the E pathway hypothesis and for a crucial functional role of Glu-C180. |
==About this Structure== | ==About this Structure== | ||
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[[Category: Succinate dehydrogenase]] | [[Category: Succinate dehydrogenase]] | ||
[[Category: Wolinella succinogenes]] | [[Category: Wolinella succinogenes]] | ||
| - | [[Category: Lancaster, C | + | [[Category: Lancaster, C R.D.]] |
[[Category: CIT]] | [[Category: CIT]] | ||
[[Category: F3S]] | [[Category: F3S]] | ||
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[[Category: tricarboxylic acid cycle]] | [[Category: tricarboxylic acid cycle]] | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:41:03 2008'' |
Revision as of 14:41, 21 February 2008
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GLU C180-> GLN VARIANT QUINOL:FUMARATE REDUCTASE FROM WOLINELLA SUCCINOGENES
Overview
Reconciliation of apparently contradictory experimental results obtained on the quinol:fumarate reductase, a diheme-containing respiratory membrane protein complex from Wolinella succinogenes, was previously obtained by the proposal of the so-called "E pathway hypothesis." According to this hypothesis, transmembrane electron transfer via the heme groups is strictly coupled to cotransfer of protons via a transiently established pathway thought to contain the side chain of residue Glu-C180 as the most prominent component. Here we demonstrate that, after replacement of Glu-C180 with Gln or Ile by site-directed mutagenesis, the resulting mutants are unable to grow on fumarate, and the membrane-bound variant enzymes lack quinol oxidation activity. Upon solubilization, however, the purified enzymes display approximately 1/10 of the specific quinol oxidation activity of the wild-type enzyme and unchanged quinol Michaelis constants, K(m). The refined x-ray crystal structures at 2.19 A and 2.76 A resolution, respectively, rule out major structural changes to account for these experimental observations. Changes in the oxidation-reduction heme midpoint potential allow the conclusion that deprotonation of Glu-C180 in the wild-type enzyme facilitates the reoxidation of the reduced high-potential heme. Comparison of solvent isotope effects indicates that a rate-limiting proton transfer step in the wild-type enzyme is lost in the Glu-C180 --> Gln variant. The results provide experimental evidence for the validity of the E pathway hypothesis and for a crucial functional role of Glu-C180.
About this Structure
2BS3 is a Protein complex structure of sequences from Wolinella succinogenes with , , , , , , and as ligands. Active as Succinate dehydrogenase, with EC number 1.3.99.1 Known structural/functional Site: . Full crystallographic information is available from OCA.
Reference
Experimental support for the "E pathway hypothesis" of coupled transmembrane e- and H+ transfer in dihemic quinol:fumarate reductase., Lancaster CR, Sauer US, Gross R, Haas AH, Graf J, Schwalbe H, Mantele W, Simon J, Madej MG, Proc Natl Acad Sci U S A. 2005 Dec 27;102(52):18860-5. PMID:16380425
Page seeded by OCA on Thu Feb 21 16:41:03 2008
Categories: Protein complex | Succinate dehydrogenase | Wolinella succinogenes | Lancaster, C R.D. | CIT | F3S | FAD | FES | HEM | LMT | NA | SF4 | 2fe-2s | 3d-structure | 3fe-4s | 4fe-4s | Citric acid cycle | Dihaem cytochrome b | Electron transport | Fad | Flavoprotein | Fumarate reductase | Heme | Ion-sulphur protein | Iron | Iron-sulfur | Metal-binding | Oxidoreductase | Respiratory chain | Transmembrane | Tricarboxylic acid cycle
