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Discovery of General Controů Nondereprssible 5 protein interactors using proximity labeling
Potapenkova, Uliana
Usually, proteins don’t act as isolated species, but they perform their function in complexes with other proteins. Identification of protein-protein interactions (PPIs) brings insights into regulation of plant developmental processes and their response to the environmental conditions. In recent years, several methods to study PPIs were developed, since they represent the key objectives of plant system biology research. This thesis is focused on the Proximity Labeling (PL) assay in order to study GCN5 protein interactors. GCN5 is a histone acetyltransferase that plays a crucial role in plants’ defense responses to various abiotic stresses. Studies of its interactors provide valuable information on the histone acetylation mechanisms and the way plants cope with adverse conditions. Here, the advantages and disadvantages of PL assay are summarized and compared with other methods of PPIs studies. In the experimental part of this work, GCN5 was cloned and plasmid for further biotin-based proximity labeling was prepared.
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Current methods in protein-protein interactions research
Křivková, Jana ; Hrdý, Ivan (advisor) ; Kučerová, Jitka (referee)
Protein-protein interactions (PPI) have a crucial role in all processes in living cells. Understanding the interactions between proteins allows us to describe cell processes in more detail and their study opens new possibilities for drug design. The importance of the question of studying PPI is shown in the recent development of various methods for their identification and description. The aim of this thesis is to give an overview of new and improved experimental methods of identification and characterization of protein-protein interactions. Methods described in this thesis are divided in four chapters - proximity-dependent labelling methods (BioID, BioID2, APEX, TurboID, MiniTurbo, PUP-IT, AirID, SPPLAT, EMARS), cross-linking methods (XS-MS), fluorescence methods for identification and visualization (BiFC, FRET, BRET) and biophysical methods for description of kinetics and thermodynamics parameters of interaction (SPR, ITC, MT).
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