The structure of an engineered domain-swapped ribonuclease dimer and its implications for the evolution of proteins toward oligomerization

Background: Domain swapping has been proposed as a mechanism that explains the evolution from monomeric to oligomeric proteins. Bovine and human pancr eatic ribonucleases are monomers with no biological properties other than t heir RNA cleavage ability. In contrast, the closely related bovine seminal ribonuclease is a natural domain-swapped dimer that has special biological properties, such as cytotoxicity to tumour cells. Several recombinant ribon uclease variants are domain-swapped dimers, but a structure of this kind ha s not yet been reported for the human enzyme. Results: The crystal structure at 2 Angstrom resolution of an engineered ri bonuclease variant called PM8 reveals a new kind of domain-swapped dimer, b ased on the change of N-terminal domains between the two subunits. The swap ping is fastened at both hinge peptides by the newly introduced Gln101, inv olved in two intermolecular hydrogen bonds and in a stacking interaction be tween residues of different chains. Two antiparallel salt bridges and water -mediated hydrogen bonds complete a new interface between subunits, while t he hinge loop becomes organized in a 3(10) helix structure. Conclusions: Proteins capable of domain swapping may quickly evolve toward an oligomeric form. As shown in the present structure, a single residue sub stitution reinforces the quaternary structure by forming an open interface. An evolutionary advantage derived from the new oligomeric state will fix t he mutation and favour others, leading to a more extended complementary dim erization surface, until domain swapping is no longer necessary for dimer f ormation. The newly engineered swapped dimer reported here follows this hyp othetical pathway for the rapid evolution of proteins.