ИСТИНА

Introduction: The protein folding to a native functional structure is the key question of protein science. At present, our understanding of protein folding with β-barrels remains limited. Odorant-binding proteins (OBPs) are promising building blocks for construction of optical biosensors for dangerous substances (toxic, explosives and so on). Classical OBPs share a monomer nine-stranded β-barrel structure which encloses a ligand binding site. The dimeric bovine OBP (bOBP) has a unique folding pattern with the α-helical domain of each unit crossing over the β-barrel of the other unit. Such domain swapping mechanism was observed in many proteins and has a great effect on protein structure and function. Purpose: Here we studied the folding of β-barrel proteins and the contribution of the domain swapping to their stability and unfolding – refolding probing by recombinant bOBP. The obtained results are compared with the stability of the native tissue extracted bOBP and its mutant form not liable dimerization. Materials and Methods: The main methods used are protein intrinsic fluorescence and size-exclusion chromatography. Results: The recombinant bOBP native dimer is only formed at elevated denaturant concentrations. This process requires the re-organizing the protein structure by progressing through the formation of an intermediate state. The bOBP dimerization process occurring before the protein unfolds appears to be irreversible. The domain swapping imposes stabilizing effect as revealed by experiments with bOBP mutant forms. Conclusions: The recombinant bOBP structural changes at pre-denaturing region bear a local character with overall structure of the protein being reserved; still they should be considered where the protein is used as sensitive element in a biosensor system. Despite the complex unfolding rout, bOBP is stable enough for being used in biosensor systems.