Retroevolution of the structure of lambda Cro

Abstract

The Cro protein from bacteriophage λ has a dimeric mixed α+β fold which evolved from an ancestral monomeric all-α fold. Monomeric mixed α+β structures may have been intermediates in this process. A monomeric variant of λ Cro incorporating A33W and F58D mutations has been identified and characterized. These mutations involve large changes in side chain size, and were predicted by modeling to disrupt the dimer interface while increasing the thermodynamic stability of the monomer. This combination of mutations does lower the λ Cro dissociation constant by a factor of at least 1,000, while maintaining the protein's secondary structure content. The A33W and F58D mutations also increase the maximal monomer thermal stability of λ Cro by approximately 12 degrees. The A33W/F58D Cro variant shows the plausibility of monomeric mixed α+β intermediates in the evolution of λ Cro, and shows that mutations at these positions could have been critical for evolution of the dimer.