The swapping process in bovine pancreatic ribonuclease: substitution of Pro114

The stability of the folded structure of a protein molecule is generally limited to a narrow range of environmental parameters. Depending on temperature, pH and types of co-solutes and on the evolutionary history of the protein itself, the tertiary structure may become less tightly bound and undergo to a more or less extensive unfolding that can lead to either unspecific aggregation or ordered adducts with other molecules or even to oligomeric aggregates endorsed with new functions. In the same way, specific sequence modification in hot points of the protein may lead to an “open” monomeric species which can interact in a complementary fashion with a similarly opened molecule (domain swapping) to produce a thermodynamic stable or kinetically trapped transient dimeric form (swapped dimer)[1]. The dimer is stabilized by the pre-existing surfaces of the swapped domains, whereas the flexible loops connecting these domains give rise to a somewhat ‘destructured quaternary assembly’ of the chains that strongly increases the potentiality for higher aggregation [2]. Bovine pancreatic ribonuclease (RNase A) is able to swap both the C-terminal - strand (residues 116-124) and the N-terminal -helix (residues 1-13), thus forming two different dimers [3-4]. Cis-trans isomerization of the Asn113-Pro114 peptide group was observed when the protein forms the C-terminal swapped dimer [4]. To study the effect of the substitution of Pro114 on the swapping process of RNase A, we have prepared and characterized the P114A monomeric and dimeric variants of the enzyme. Surprisingly, the mutation not only affects the swapping of the C-terminal b-strand but also that of the N-terminal helix and the stability of the two swapped dimers. These results, which are discussed on the bases of the X-ray structures of the monomeric and C-terminal swapped dimeric forms of P114A and on the available biochemical and structural data on RNase A dimers, reveal a cross-talk between the chain termini in the swapping process of RNase A.