Reciprocal domain evolution within a transactivator in a restricted sequence space
Por:
Juárez K., Flores H., Dávila S., Olvera L., González V., Morett E.
Publicada:
1 ene 2000
Resumen:
Although the concept of domain merging and shuffling as a major force in protein evolution is well established, it has been difficult to demonstrate how domains coadapt. Here we show evidence of coevolution of the Sinorhizobium meliloti NifA (SmNifA) domains. We found that, because of the lack of a conserved glycine in its DNA-binding domain, this transactivator protein interacts weakly with the enhancers. This defect, however, was compensated by evolving a highly efficient activation domain that, contrasting to Bradyrhizobium japonicum NifA (BjNifA), can activate in trans. To explore paths that lead to this enhanced activity, we mutagenized BjNifA. After three cycles of mutagenesis and selection, a highly active derivative was obtained. Strikingly, all mutations changed to amino acids already present in SmNifA. Our artificial process thus recreated the natural evolution followed by this protein and suggests that NifA is trapped in a restricted sequence space with very limited solutions for higher activity by point mutation.
Filiaciones:
Juárez K.:
Instituto de Biotecnología, Univ. Nac. Autónome de Mex., AP 510-3, Cuernavaca, Morelos, 62250, Mexico
Flores H.:
Instituto de Biotecnología, Univ. Nac. Autónome de Mex., AP 510-3, Cuernavaca, Morelos, 62250, Mexico
Dávila S.:
Instituto de Biotecnología, Univ. Nac. Autónome de Mex., AP 510-3, Cuernavaca, Morelos, 62250, Mexico
Institute de Ecología, Univ. Nac. Auton. de México, Ciudad Universitaria, 04510, Mexico, D.F., Mexico
Olvera L.:
Instituto de Biotecnología, Univ. Nac. Autónome de Mex., AP 510-3, Cuernavaca, Morelos, 62250, Mexico
González V.:
Instituto de Biotecnología, Univ. Nac. Autónome de Mex., AP 510-3, Cuernavaca, Morelos, 62250, Mexico
Ctro. Invest. Sobre Fijacion N., Univ. Nac. Auton. de México, Cuernavaca, Morelos, 62210, Mexico
Morett E.:
Instituto de Biotecnología, Univ. Nac. Autónome de Mex., AP 510-3, Cuernavaca, Morelos, 62250, Mexico
Europ. Molecular Biology Laboratory, Meyerhofstrasse 1, 69117, Heidelberg, Germany
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