Login
- Original title:
- Röntgenstrukturanalyse der ba3 Cytochrom-c Oxidase aus Thermus thermophilus und ihres Substrates Cytochrom-c552
- Translated title:
- Crystal Structure Analysis of the ba3 Cytochrome-c Oxidase from Thermus thermophilus and its Substrate Cytochrome-c552
- Author:
- Than, Manuel E.
- Year:
- 2000
- Document type:
- Dissertation
- Faculty/School:
- Fakultät für Chemie
- Advisor:
- Huber, Robert (Prof. Dr.)
- Referee:
- Michel-Beyerle, Maria-Elisabeth (Prof. Dr.); Bacher, Adelbert (Prof. Dr. Dr.)
- Format:
- Text
- Language:
- de
- Subject group:
- CHE Chemie; PHY Physik
- Translated keywords:
- Cytochrome-c Oxidase; Cytochrome-c552; Thermus thermophilus; Electron Transfer; Proton Pump; Crystal Structure; X-ray Crystallography; Membrane Protein; Crystal Transformation; Cluster Phasing
- Controlled terms:
- Thermus thermophilus ; Cytochromoxidase ; Röntgenstrukturanalyse ; Cytochrom c-552
- TUM classification:
- CHE 887d ; CHE 829d ; CHE 806d ; CHE 802d ; CHE 342d ; PHY 606d
- Abstract:
- Die dreidimensionalen Strukturen der ba3 Cytocrom-c Oxidase aus dem Eubakterium Thermus thermophilus sowie ihres Substrates Cytochrom-c552 wurden mit den Methoden der Röntgenstrukturanalyse bei einer Auflösung von 2,4 Å bzw. 1,28 Å aufgeklärt. Die Analyse dieser Strukturen ermöglichte das Verständnis der besonderen biochemischen Eigenschaften dieser beiden Proteine bezüglich Thermostabilität, Protonenpumpaktivität, Reaktionsmechanismus und Elektronenübertragung im Vergleich zu typischen Vertret... »
- Translated abstract:
- Cytochrome-c oxidases are integral membrane proteins which play a central role in the energy metabolism of aerobic organisms. They catalyze the last step in the electron transport chain, the reduction of molecular oxygen to water. The energy is thereby conserved as a transmembrane potential which in turn is utilized for the synthesis of adenosine triphosphate (ATP), the central energy carrier molecule in biological systems.
The three-dimensional structure of the ba3 cytochrome-c oxidase from Thermus thermophilus was determined at a resolution of 2.4 Å using X-ray crystallography. It is the first crystal structure of a membrane protein for which Multiple Anomalous Dispersion (MAD) was used to determine the protein phases. The structure reveales the presence of a third, new subunit. The structural understanding regarding the function and the mechanism of this enzyme was completed by solving the crystal structure of the electron donor of the ba3 cytochrome-c oxidase, the soluble cytochrome-c552, at a resolution of 1.28 Å.
A structure based sequence alignment between the phylogenetically very distant ba3-oxidase and the structurally known cytochrome-c oxidases from Paracoccus denitrificans and bovine heart leads to the elucidation of sequence motivs and structural details that seem to be essential for the function of all heme-copper-oxidases. A new, potential intramolecular electron transfer pathway leading directly from the CuA-centre to the CuB-atom could be identified. An accumulation of water molecules which is conserved in all structurally known cytochrome-c oxidases and located above the heme-propionates (water pool) most likely functions as primary acceptor of both, pumped protons and water molecules formed at the active site. Specific features of the ba3-oxidase that are required for the functionality of this enzyme under elevated temperatures and under low oxygen availability include the presence of an extended, hydrophobically coated oxygen channel which leads from the surface of molecule near the middle of the membrane to the active site as well as modifications in the proton pathways. The ba3-oxidase contains besides the classical K- and D-proton pathways the additional, potential Q-pathway. The binary, intermolecular electron transfer complex between cytochrome-c552 and the ba3-oxidase is stabilized in contrast to other members of this class of enzymes mainly by hydrophobic but not electrostatic interactions. Further aspects regarding the proton pump activity of this and other terminal heme-copper-oxidases are discussed.
The structure determination of cytochrome-c552 resulted in the identification of a structural variation of the cytochrome-c fold which is responsible for the thermostability of this protein, a C-terminal clamp that tightly encloses the rest of the molecule. The high hydrophobicity and compactness of the protein interior contribute further to the enhanced stability.
The characterization of the trasformation behavior of the ba3-oxidase crystals was essential for the succesful determination of its crystal structure and resulted in the development of methods which are of great interest for many protein crystals. The ba3-oxidase crystals proceed through several preferred crystal states when they are subjected to a tightly controlled dehydration. This procedure leads to an extensive increase in diffraction power in comparison to the native crystals. Successful shock freezing of the crystals which prevents radiation damage during data collection is only possible with crystals that were transformed by dehydration to a crystal state with lower solvent content.
The determination of the crystal structure of the periplasmatic nitrate reductase from Desulfovibrio desulfuricans was only possible after solving the exact coordinates of the four iron atoms of its [4Fe-4S]-cluster using the known geometry of this cluster and a procedure based on patterson correlation refinement. This method together with the collected MAD-data made the determination of high quality protein phases at high resolution possible.
During the course of this work, the crystal structure of the first biliprotein-linker-complex could be solved, which resulted from overcoming statistical disorder due to crystallization in a new space group. - Publication :
- Universitätsbibliothek der TU München
- WWW:
- https://mediatum.ub.tum.de/?id=601138
- Date of submission:
- 10.04.2000
- Oral examination:
- 23.05.2000
- File size:
- 6164718 bytes
- Pages:
- 156
- Urn (citeable URL):
- https://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:91-diss2000052306175
- Last change:
- 05.06.2007
BibTeX