title: 3D domain swapping provides a minor alternative refolding pathway for ribonuclease A.
abstract: The C-terminal β-hairpin of RNase A contains a turn with a cis Asn113-Pro114 peptide bond. Pioneering pulsed HX experiments have shown that the C-terminal β-hairpin forms early during refolding. This is puzzling since the Asn113-Pro114 bond is predominately trans at this stage and this conformation destabilizes the native monomer. RNase A, when refolded at high concentration, forms a series of 3D domain-swapped oligomers. In the oligomers formed by C-terminal β-strand swapping, Asn113-Pro114 is trans and permits the formation of a new intersubunit β-sheet. We hypothesize that oligomeric species with trans Asn113-Pro114 may form during refolding. Such species could account for the HX results while comfortably accommodating Asn113-Pro114 in the trans conformation. Here, we test this hypothesis by employing chromatographic methods to detect oligomers forming in refolding conditions and find significant amounts of dimer. We propose that a 3D domain-swapped dimeric intermediate provides a minor alternative pathway for RNase A refolding.
authors.affiliation: Dipartimento di Chimica, Università degli Studi di Napoli Federico II, Napoli, Italy.
authors.forename: Carmine
authors.initials: C
authors.lastname: Ercole
authors.forename: Douglas V
authors.initials: DV
authors.lastname: Laurents
mesh_terms: D002845:Chromatography; D019281:Dimerization; D011489:Protein Denaturation; D058849:Protein Refolding; D017434:Protein Structure, Tertiary; D012259:Ribonuclease, Pancreatic / Q000737:chemistry*
chemical_list: D012259:Ribonuclease, Pancreatic