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Identifier

000371503

Title

Απομόνωση και δομικός χαρακτηρισμός υποσυμπλόκων και πρωτεϊνών του πυρήνα του φωτοσυστήματος ΙΙ των ανώτερων φυτών

Alternative Title

Isolation and structural characterization of photosystem II core subcomplexes and proteins from higher plants

Author

Ψυλινάκης, Εμμανουήλ Αντ.

Thesis advisor

Γανωτάκης, Δημήτριος

Abstract

Photosystem II is a membrane multiprotein complex, which catalyzes water splitting, via a unique mechanism. The significant functional properties of this complex are attributed to its unique structural characteristics. Indirect biochemical studies have yielded information on the location of the PSII proteins. Recently, electron microscopy studies have led to models concerning the relative assembly and subunit topology in PSII. However, the only available information on the PSII structure on a molecular level derive from purple photosynthetic bacteria, whose structure has been determined by X-ray crystallography. To date, a three-dimensional model, which would reveal the unique functional properties of PSII, is absent. In the present work the aim has been the structural study of PSII on a molecular level, as well as the level of PSII assembly. The study of the molecular structure of PSII was based on the production of three dimensional crystals and the analysis of the structure with X-ray crystallography. The analysis of the tertiary structure of PSII was based on electron microscopy studies of the reaction center. Concerning the crystallization of PSII, a new purification protocol was established, which enabled quantitative isolation of CP 47 and the reaction center complex, in a stable state. This protocol is based on the selective extraction of membrane proteins, using the appropriate combination of detergents and the enrichment of the subcomplexes with ultracentrifugation, using a linear sucrose gradient. The structural study of the PSII reaction center complex yielded information on the oligomerization state and the location of the membrane proteins in the PSII-core. The reaction center complex was isolated in a monomeric form and its molecular weight was estimated by a combination of two independent techniques. Analysis of the images, which were obtained by electron microscopy, led to a protein density map of the D1 and D2 subunits. A comparison of the reaction center map to the map of the whole PSII-core complex revealed the relative positions of the major membrane proteins. These results supported a central location for the D1/D2 heterodimer and a peripheral location for CP 47 and CP 43, giving the complex a pseudo twofold symmetry.

Language

Greek

Subject

Photosynthesis

Ανώτερα φυτά

Μεμβρανικές πρωτεϊνες

Φωτοσύνθεση

Φωτοσύστημα ΙΙ

Issue date

2011-11-01

Collection

School/Department--School of Sciences and Engineering--Department of Chemistry--Post-graduate theses