exploring the structure and function of cytochrome bo 3 ubiquinol oxidase from escherichia coli
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Exploring the Structure and Function of Cytochrome bo 3 Ubiquinol Oxidase from Escherichia coli. Lai Lai Yap Department of Biochemistry. Heme-Copper Oxidase Superfamily. catalyze reduction of oxygen to water , and utilizes free energy produced to pump protons across membrane - PowerPoint PPT PresentationTRANSCRIPT
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Exploring the Structure and Function of Cytochrome bo3 Ubiquinol Oxidase from
Escherichia coli
Lai Lai Yap
Department of Biochemistry
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Heme-Copper Oxidase Superfamily
• catalyze reduction of oxygen to water, and utilizes free energy produced to pump protons across membrane
• transmembrane proton and voltage gradient thus generated is converted to useful energy forms (eg. ATP by ATP synthase)
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• 2 main groups (based on electron-donating substrate) : cytochrome c oxidase (eg. mitochondrial cytochrome c oxidase) and ubiquinol oxidase (eg. cytochrome bo3 ubiquinol oxidase)
• membership based on presence of subunit homologous to subunit I of mammalian cytochrome c oxidase
• subunit I:
– largest subunit
– binuclear center (where O2 binds and is reduced to water) consisting of a heme and copper (CuB)
– a second heme (which facilitates transfer of electrons to binuclear center)
Heme-Copper Oxidase Superfamily
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Respiratory Chains of E. coli
High O2 affinity
Low O2 affinitySubstrates eg. NADH, succinate
Dehydrogenases
Quinone
Cytochrome bo3
Cytochrome bd
O2
O2
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Cytochrome bo3 ubiquinol oxidase
• a terminal oxidase in the aerobic respiratory chain of Escherichia coli
• member of the heme-copper oxidase superfamily
• consists of four subunits
• catalyzes two-electron oxidation of ubiquinol-8 (Q8H2) at periplasmic side of cytoplasmic membrane and four-electron reduction of oxygen to water at cytoplasmic side
• also functions as a proton pump by translocating protons across the cytoplasmic membrane to establish an electrochemical proton gradient
• possible mechanism: ubiquinone bound at the high affinity site (QH) acts as cofactor and mediates electron transfer from ubiquinol substrate (at the low-affinity QL site) to heme b
• reduced heme b then provides electrons to the binuclear center for the reduction of oxygen to water
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Electron and proton transfer in cytochrome bo3 ubiquinol oxidase
QL
QH
II IIII IV
½ O2 + 2H+H2O 2H+
translocation
2H+
2e-2e-
2e-
2e-
periplasm
cytoplasm
Heme b
Heme o3
CuB
QH2
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Subunit ISubunit IISubunit IIISubunit IV
P
C
ubiquinol binding site
Spherical rendering structure of ubiquinol oxidase
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P
C
ubiquinol binding site
Subunit ISubunit IISubunit IIISubunit IV
Structure of ubiquinol oxidase
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Heme b Heme o3
CuB
His106
His421
His334
His333
His284
His419
Redox metal centers of ubiquinol oxidase (in subunit I)
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Rainbow rendering of ubiquinol oxidase and cytochrome c oxidase
Cytochrome c oxidase Ubiquinol oxidase
Cytochrome c binding site Ubiquinol binding site
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Superposition of ubiquinol oxidase and cytochrome c oxidase
Ubiquinol oxidase
Cytochrome c oxidase
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Proton transfer pathways
• D- and K-channels in subunit I
• channels form polar cavities that originate on the cytoplasmic side, leading to the binuclear center for proton pumping and water formation
• D-channel : uptake of both chemical and pumped protons
• K-channel : load enzyme with protons at some earlier catalytic steps
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D- and K-channels of ubiquinol oxidase
D-channel K-channelD-channel K-channel
D135T211
N142
N124
S145
T204
T149
T201
E286
H106
H421
H333
H334
H284
Y288
T359
K362
S315S299
H+ out
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QL
QH
II IIII IV
½ O2 + 2H+H2O 2H+
translocation
2H+
2e-2e-
2e-
2e-
periplasm
cytoplasm
Heme b
Heme o3
CuB
QH2
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P
C
ubiquinol binding site
Subunit ISubunit IISubunit IIISubunit IV
Structure of ubiquinol oxidase
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Ubiquinol oxidase with modeled ubiquinone (at ubiquinol binding site)
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ubiquinone
L160
R71
D75
M78
M79I102
H98
Q101
Ubiquinone binding site of ubiquinol oxidase with modeled ubiquinone
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Electron and proton transfer in cytochrome bo3 ubiquinol oxidase
QL
QH
II IIII IV
½ O2 + 2H+H2O 2H+
translocation
2H+
2e-2e-
2e-
2e-
periplasm
cytoplasm
Heme b
Heme o3
CuB
QH2
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ubiquinone
heme b
heme o3
CuB
M79I102
H106
H421
H419
Membrane normal view of subunit I with modeled ubiquinone
possible electron path from ubiquinone to binuclear center