Analysen zu Funktionen von zwei Vertretern der Oxa1/Alb3/YidC-Proteinfamilie

Abstract

The aim of this PhD thesis was the functional analysis of Oxa1/Alb3/YidC homologues in Synechocystis and Chlamydomonas reinhardtii with focus on protein insertion and assembly. The results of the analysis of Synechocystis Slr1471p using the Split-Ubiquitin system can be summarized as follows: A direct interaction of the PSI reaction centrum protein PsaB with the Oxa1/Alb3/YidC homologue Slr1471p and the cpSecY homologue Sll1814p could be demonstrated. Localization studies of the interaction sites of Slr1471p showed that all analyzed proteins interact with the transmembrane helices 4-5 of Slr1471p, apart from Slr1531p. Slr1531p associates with the N-terminal transmembrane helices 1-3 of Slr1471p. This suggests that the N-terminal site of Slr1471p binds to Slr1531p and is responsible for protein targeting to the membran, while the C-terminal site shows an affinity for the substrates. For the biochemical analysis of Alb3.2p of C. reinhardtii the following results were obtained: Alb3.2p is localized in the thylakoid membrane and enriched in the stroma lamella. An association of Alb3.2p with polysomes could not be demonstrated using polysome analysis. Therefore an Alb3.2p participitation in co-translational integration is very unlikely. Alb3.2p is interacting only with itself, as shown by co-immunoprecipitation and HPLC/MS analysis. In the chloroplasts of C. reinhardtii Alb3.2p exist as monomer or homooligomer. An association of Alb3.2p with interacting partners seems to be weak or short-lived. An association of Alb3.2p with high molecular weight complexes of > 1 MDa could be demonstrated for the first time by cross linking experiments. The results give evidence that the function of Oxa1/Alb3/YidC homologues changed during evolution. Because of gene duplication it was possible that Oxa1/Alb3/YidC homologues in the chloroplasts of higher plants gained spezialized functions loosing the property to insert proteins co-translationally.