It is well-established in the literature that the structure and stability of proteins is influenced by co-solutes and other nearby structures. Recent reports on molecular crowding and confinement demonstrate that the effects of a protein’s immediate environment are complex and involve several different phenomena, such as the protein’s conformational entropy, changes in the properties of confined solvent molecules and enthalpic interactions. Changes in the secondary and tertiary levels of structure upon crowding were explored using a novel protein system. Osteocalcin is a small, three-helix protein with minimal tertiary structure. Jak-1 is a peptide that forms a single alpha helix with no tertiary structure. Both proteins are relatively disordered in dilute solution, and both transition to become more folded upon binding of calcium to their three gamma-carboxylated glutamic acid residues. This unique system permits direct comparison between a protein with and a protein without tertiary structure. We will describe our observations of the effects on structure and stability of osteocalcin and Jak-1 upon encapsulation in sol-gels, crowding with Ficoll-70 and crowding with the small molecule osmolyte TMAO. In brief, secondary structure stabilization was favored over tertiary structure stabilization for sol-gel encapsulation, while the opposite was true for osmolyte crowding with TMAO. Furthermore, we will discuss these results in the context of the current knowledge in the field of protein molecular crowding.

Use of a unique protein model system to explore the effects of crowding by sol-gel confinement, polymeric crowding and small-molecule osmolyte crowding on different levels of protein structure

251st National Meeting (2016)

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