From genomics to proteomics in Azoarcus sp. BH72 a N2-fixing endophytic bacterium

Abstract

Since the first genome sequence was completed in 1995, the number of publicly available prokaryotic genomes increased exponentially up today, when more than 300 microbial projects had been completed. Within this 'genomic revolution', the complete genome analysis of environmentally and biotechnologically relevant microorganisms constitutes an emerging field of research. In this context the genome-sequencing project of Azoarcus sp. BH72 took place. The diazotrophic endophyte Azoarcus sp. BH72 posses a high biotechnological potential as it is capable of colonizing the interior of rice roots, which is one of the globally most important crops. Moreover, since the rice genome is also sequenced, the Azoarcus-rice system represents a great model for bacteria-grass interaction studies. In the present thesis three main topics, from genomics to functional genomic studies, were covered. The aim of the first part of this work was the characterization of an Azoarcus sp. BH72 bacterial artificial chromosome (BAC) library as well as the building of a physical map of strain BH72 chromosome. Both tools were used for an independent analysis of the genome structure in comparison to a shotgun library with small insert sizes, for contig assembly and gap closure of this shotgun library and for genome comparison analysis. The second main objective of this work was the annotation and analysis of a part of the genome sequence. In particular, genes which belong to the COG categories 'Ion Transporters and Metabolism', and 'Carbohydrates Transport and Metabolism', were studied. This analysis revealed several highlights in the genome sequence, particularly in the iron metabolism, which can be used as a starting point in future studies. Finally, in the last part of this work a functional genomic analysis of Azoarcus sp. BH72 grown under different conditions of N2-fixation was performed, using a differential-display proteomic approach. Proteomic patterns of strain BH72 N2-fixing cells, in pure- and co-culture with the endophytic fungus Acremonium alternatum, showed strongly significant differences. The identification of the major proteins showed that the nitrogen metabolism was very active in both conditions, as well as the carbon metabolism, which was adapted to the carbon sources available. Several membrane proteins were identified which most probably are involved in bacteria-fungus interaction, as well as in bacterial response to fungus metabolites. The results obtained contribute to a better understanding of the Azoarcus sp. BH72 physiology and ecology.