Extensive modifications for methionine enhancement in the beta-barrels do not alter the structural stability of the bean seed storage protein phaseolin.
J Protein Chem
; 14(8): 665-78, 1995 Nov.
Article
em En
| MEDLINE
| ID: mdl-8747427
ABSTRACT
Common beans are widely utilized as a food source, yet are low in the essential amino acid methionine. As an initial step to overcome this defect the methionine content of the primary bean seed storage protein phaseolin was increased by replacing 20 evolutionarily variant hydrophobic residues with methionine and inserting short, methionine-rich sequences into turn and loop regions of the protein structure. Methionine enhancement ranged from 5 to 30 residues. An Escherichia coli expression system was developed to characterize the structural stability of the mutant proteins. Proteins of expected sizes were obtained for all constructs except for negative controls, which were rapidly degraded in E. coli. Thermal denaturation of the purified proteins demonstrated that both wild-type and mutant phaseolin proteins denatured reversibly at approximately 61 degrees C. In addition, urea denaturation experiments of the wild-type and a mutant protein (with 30 additional methionines) confirmed that the structural stability of the proteins was very similar. Remarkably, these results indicate that the phaseolin protein tolerates extensive modifications, including 20 substitutions and two loop inserts for methionine enhancement in the beta-barrel and loop structures, with extremely small effects on protein stability.
Buscar no Google
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas de Plantas
/
Conformação Proteica
/
Engenharia de Proteínas
/
Metionina
Idioma:
En
Revista:
J Protein Chem
Ano de publicação:
1995
Tipo de documento:
Article
País de afiliação:
Estados Unidos