Non covalent coupling of proteins to a oligomeric scaffold through high affinity heterodimeric modules

VIGNALE F; GOROJOVSKY N; SANTOS J; GOLDBAUM FA; CRAIG PO

Salto

Congreso; Latin American Crosstalk in Biophysics and Physiology (SBF.uy‐SAB/2015); 2015

Seccional Biofísica de la Sociedad Uruguaya de Biociencias (SBF.uySUB), Sociedad Argentina de Biofisica (SAB) y LAFeBS

The polymeric display of molecules is an emerging technology for many applications. It could be used to increase the immunogenicity of antigens, the interaction strength of protein domains for polymeric ligands, and the production of multifunctional particles. Brucella Abortus Lumazine Synthase (BLS) is a decameric protein composed of a 17 kDa subunit arranged as a dimer of pentamers. In this work, we describe the use of BLS as a oligomeric scaffold for the polymeric display of proteins. For this aim we developed a non-covalent coupling method based on the use of high affinity heterodimeric modules complementary fused to the structure of BLS and target proteins. Results obtained with a variety of protein models (9 kDa double stranded RNA binding domain from murine Staufen-1 protein, 18 kDa sialic acid binding domain from Bovine Rotavius VP4 capsid protein, and 26 kDa jellyfish Green Fluorescent protein) indicate that is possible to obtain well folded and stable decameric particles by this method, as indicated by the analysis of these constructions though CD, fluorescence, and light scattering techniques. The immunological use of BLS as a carrier of antigens is illustrated with the RBD3 antigen. These results demonstrates the potential of the BLS carrier approach to deliver antigens to the immune system in a highly order array, preserving their natural conformation and enhancing their immunogenicity. In addition, an increase in the interaction strength of RBD3 for a model double stranded RNA ligand could also be observed when this domain was coupled to BLS. We present valuable data for the design, production and characterization of macromolecular assemblies and multidomain proteins for its use in the bionanotechnology field.