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Identifier

000410408

Title

Συγκριτική πρωτεομική ανάλυση ετεροκυστών από διαφορετικά στελέχη της Anabaena sp. PCC 7120

Alternative Title

Comparative proteomic analysis of heterocysts from different strains of Anabaena sp. PCC 7120

Author

Κουρπά, Αικατερίνη

Thesis advisor

Τσιώτης, Γεώργιος

Reviewer

Γανωτάκης, Δημήτριος
Κοτζαμπάσης, Κυριάκος

Abstract

Anabaena sp. PCC 7120 is a Gram negative cyanobacterium which has the ability to perform both oxygenic photosynthesis and nitrogen fixation. One of the main characteristics of this bacterium is that it is able to spatially separate these procedures by growing two different cell types (vegetative cells and heterocysts). Cell differentiation was performed by growing the bacterium in BG11 medium for about 10 days and then in BG11⁰ for 48 hours. In the present study, a protocol for the isolation and dissolution of heterocysts was applied for the three strains, WT, ΔHupL, ΔHoxH and two different methods for their proteomic analysis were tested. In the first method, the heterocysts proteins were separated by 2D IEF-SDS-PAGE electrophoresis, the peptides were obtained with in gel-digestion and they were identified by MALDI-TOF/TOF MS/MS. A total of 125 different proteins were identified, including important proteins which are involved in the photosynthesis process and in the response of the cell to stress conditions. Also, proteins that take place in the differentiation of heterocysts and nitrogen fixation, were identified, with a greater amount of them in the wild type. More generally, several unique proteins were identified for each strain, while it is interesting to note that for the ΔHoxH strain more proteins were found in total. In the second approach, the solubilized heterocysts were processed by the FASP method and the harvested peptides were identified by nLC-ESI-MS / MS. In total, 2728 different proteins were identified for all three strains, with a higher percentage in the wild type. Among other, important proteins associated with nitrogen fixation and differentiation of vegetative cells into heterocysts were found in all three strains. This process, in addition to the larger number of identified proteins, allowed us to identify much more membrane proteins compared to the first approach. In addition, it provided interesting results on the function of uptake and bidirectional hydrogenases in both wild type and mutant strains as well as the possibility of investigating the effect of genetic modifications on the function of both nitrogenase and hydrogenases in ΔHoxH and ΔHupL . More generally, both methods, but especially the second one, allowed us to estimate that the isolation and dissolution of the heterocysts were achieved to a satisfactory degree, offering a better understanding of their metabolism and function.

Language

Greek

Subject

FASP

IEF-SDS-PAGE 2D

nLC-ESI-MS/MS

Ετεροκύστες

Issue date

2017-07-21

Collection

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

Type of Work--Post-graduate theses