Genetic Perturbation Alters Functional Substates in Alkaline Phosphatase.
J Am Chem Soc
; 145(5): 2806-2814, 2023 02 08.
Article
em En
| MEDLINE
| ID: mdl-36706363
ABSTRACT
Enzymes inherently exhibit molecule-to-molecule heterogeneity in their conformational and functional states, which is considered to be a key to the evolution of new functions. Single-molecule enzyme assays enable us to directly observe such multiple functional states or functional substates. Here, we quantitatively analyzed functional substates in the wild-type and 69 single-point mutants of Escherichia coli alkaline phosphatase by employing a high-throughput single-molecule assay with a femtoliter reactor array device. Interestingly, many mutant enzymes exhibited significantly heterogeneous functional substates with various types, while the wild-type enzyme showed a highly homogeneous substate. We identified a correlation between the degree of functional substates and the level of improvement in promiscuous activities. Our work provides much comprehensive evidence that the functional substates can be easily altered by mutations, and the evolution toward a new catalytic activity may involve the modulation of the functional substates.
Texto completo:
1
Coleções:
01-internacional
Contexto em Saúde:
3_ND
Base de dados:
MEDLINE
Assunto principal:
Conformação Proteica
/
Proteínas de Escherichia coli
/
Fosfatase Alcalina
/
Escherichia coli
Idioma:
En
Revista:
J Am Chem Soc
Ano de publicação:
2023
Tipo de documento:
Article