[Estrategias para seleccionar sitios de mutacion para el incremento de metionina en la proteina de almacenamiento, faseolina, en semillas de frijol].
1994
Dyer J.M. | Nelson J.W. | Murai N.
The complete three-dimensional structure of the bean seed storage phaseolin was generated from alfa-carbon coordinates by using molecular mechanic calculations. This structure was used as a template to simulate modifications aimed at increasing the methionine content of phaseolin. A hydrophilic, methionine-rich looping insert sequence was designed. Simulated mutagenesis shows that the insert might be accommodated in turn and loop regions of the protein, but not within an alfa-helix. Methionine content was also increased by the replacement of hydrophobic amino acids with methionine in the central core beta-barrels of the phaseolin protein. Calculations indicated that methionine can effectively replace conserved or variant leucine, isoleucine, and valine residues. However, alanine residues were much more sensitive to substitution, and demonstrated high variability in the effect of methionine replacement. Introduction of multiple substitutions in the barrel interior demonstrated that the replaced residues could interact favorably to relieve local perturbations caused by individual substitutions. Molecular dynamics simulations were also utilized to study the structural organization of phaseolin. The calculations indicate that there are extensive packing interactions between the major domains of phaseolin, which have important implications for protein folding and stability. Since the proposed mutant proteins can be produced and studied, the results presented here provide an ideal test to determine if there is a correlation between the effects obtained by computer simulation and the effects of the mutations on the protein structure expressed in vivo.
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