Biophysical Society Thematic Meeting| Lima 2019

Revisiting the Central Dogma of Molecular Biology at the Single-Molecule Level

Poster Abstracts

4-POS Board 4 ALLOSTERIC MODULATIONS IN SYNTHETIC FUSION PROTEINS Kristyna Bousova 1 ; Konstantinos Tripsianes 2 ; Jiri Vondrasek 1 ; 1 Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Bioinformatics, Prague, Czech Republic 2 CEITEC, Protein-DNA Interactions, Brno, Czech Republic The design of chimeric synthetic proteins utilizing individual protein domains or artificial mini- proteins as building blocks is an important advance for the formation of new functional biomolecules for medical applications and biotechnology. It is increasingly anticipated that synthetically fused domains from different protein sources, whether derived from nature or from in silico predictions, could lead to improved properties of specifically engineered proteins through allosteric modulation of fused domains. We decided to prepare two fusion molecules in their domain order: the "forward" and the "reverse" one, to study how the domains influence each other in their different orders, and whether the order in the new protein molecules is crucial. To study the domains allosteric modulations we have proposed novel two-domain molecules composed of a well structurally characterized domain of the Postsynaptic Density Protein 3 (PDZ3; part of ZO-1) and the artificial protein Tryptophan-Cage. To decipher changes in new biomolecules we used biochemical, biophysical and structural (nuclear magnetic resonance - NMR) analyses. To determine changes in binding specificities of fusions, we used a peptide derived from PDZ3 natural binding partner molecule Junctional Adhesion Molecule A (JAM-A). Based on biophysical characterizations and NMR results, we have found out that domains order in the context of the protein can be a fundamental factor for allosteric modulation and it can determine the protein-specific function.

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