dc.contributor.author
Hugentobler, Katharina Gloria
dc.contributor.author
Heinrich, Dorothea
dc.contributor.author
Berg, Johan
dc.contributor.author
Heberle, Joachim
dc.contributor.author
Brzezinski, Peter
dc.contributor.author
Schlesinger, Ramona
dc.contributor.author
Block, Stephan
dc.date.accessioned
2020-11-30T15:53:19Z
dc.date.available
2020-11-30T15:53:19Z
dc.identifier.uri
https://refubium.fu-berlin.de/handle/fub188/28979
dc.identifier.uri
http://dx.doi.org/10.17169/refubium-28729
dc.description.abstract
The transmembrane protein cytochrome c oxidase (CcO) is the terminal oxidase in the respiratory chain of many aerobic organisms and catalyzes the reduction of dioxygen to water. This process maintains an electrochemical proton gradient across the membrane hosting the oxidase. CcO is a well-established model enzyme in bioenergetics to study the proton-coupled electron transfer reactions and protonation dynamics involved in these processes. Its catalytic mechanism is subject to ongoing intense research. Previous research, however, was mainly focused on the turnover of oxygen and electrons in CcO, while studies reporting proton turnover rates of CcO, that is the rate of proton uptake by the enzyme, are scarce. Here, we reconstitute CcO from R. sphaeroides into liposomes containing a pH sensitive dye and probe changes of the pH value inside single proteoliposomes using fluorescence microscopy. CcO proton turnover rates are quantified at the single-enzyme level. In addition, we recorded the distribution of the number of functionally reconstituted CcOs across the proteoliposome population. Studies are performed using proteoliposomes made of native lipid sources, such as a crude extract of soybean lipids and the polar lipid extract of E. coli, as well as purified lipid fractions, such as phosphatidylcholine extracted from soybean lipids. It is shown that these lipid compositions have only minor effects on the CcO proton turnover rate, but can have a strong impact on the reconstitution efficiency of functionally active CcOs. In particular, our experiments indicate that efficient functional reconstitution of CcO is strongly promoted by the addition of anionic lipids like phosphatidylglycerol and cardiolipin.
en
dc.format.extent
17 Seiten
dc.rights.uri
https://creativecommons.org/licenses/by/4.0/
dc.subject
proton translocation
en
dc.subject
single molecule
en
dc.subject
electron transfer
en
dc.subject
membrane protein
en
dc.subject
single enzyme fluorescence microscopy
en
dc.subject.ddc
500 Naturwissenschaften und Mathematik::540 Chemie::540 Chemie und zugeordnete Wissenschaften
dc.subject.ddc
500 Naturwissenschaften und Mathematik::540 Chemie::541 Physikalische Chemie
dc.title
Lipid Composition Affects the Efficiency in the Functional Reconstitution of the Cytochrome c Oxidase
dc.type
Wissenschaftlicher Artikel
dcterms.bibliographicCitation.articlenumber
6981
dcterms.bibliographicCitation.doi
10.3390/ijms21196981
dcterms.bibliographicCitation.journaltitle
International Journal of Molecular Sciences
dcterms.bibliographicCitation.number
19
dcterms.bibliographicCitation.originalpublishername
MDPI
dcterms.bibliographicCitation.volume
21
dcterms.bibliographicCitation.url
https://doi.org/10.3390/ijms21196981
refubium.affiliation
Biologie, Chemie, Pharmazie
refubium.affiliation.other
Institut für Chemie und Biochemie / Organische Chemie
refubium.note.author
Die Publikation wurde aus Open Access Publikationsgeldern der Freien Universität Berlin gefördert.
refubium.resourceType.isindependentpub
no
dcterms.accessRights.openaire
open access
dcterms.isPartOf.eissn
1422-0067