Retroevolution of Cro toward a stable monomer
نویسندگان
چکیده
The Cro protein from bacteriophage has a dimeric fold that evolved from an ancestral allmonomer. The sequence mutations responsible for this dramatic structural evolution are unknown. Here we use analysis of sequence alignments to show that Ala-33, a small side chain in the hydrophobic ‘‘ball-and-socket’’ dimer interface of Cro, was a much larger tryptophan side chain at a previous point in evolution. The retroevolutionary Cro-A33W mutant shows a 10-fold reduction in dimerization affinity relative to the wild type as well as a large increase in monomer thermal stability ( Tm > 10°C), apparently due to partial filling of the hydrophobic socket from within the same monomer. An additional mutation in the dimer interface, F58D, almost completely abolishes detectable dimerization while maintaining the high monomer stability. The secondary structure content of the monomerized versions of Cro is similar to that of the wild-type protein, and the tertiary structure of the monomer appears relatively well defined. These results (i) support a model in which the ball-and-socket dimer interface of Cro was created by altered volume mutations within a limited branch of the Cro lineage and (ii) suggest the possibility that the evolution of the dimer from an allmonomer proceeded through an monomer intermediate.
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