Conserved residues of the Pro103-Arg115 loop are involved in triggering the allosteric response of the Escherichia coli ADP-glucose pyrophosphorylase.
Protein Sci
; 24(5): 714-28, 2015 May.
Article
em En
| MEDLINE
| ID: mdl-25620658
ABSTRACT
The synthesis of glycogen in bacteria and starch in plants is allosterically controlled by the production of ADP-glucose by ADP-glucose pyrophosphorylase. Using computational studies, site-directed mutagenesis, and kinetic characterization, we found a critical region for transmitting the allosteric signal in the Escherichia coli ADP-glucose pyrophosphorylase. Molecular dynamics simulations and structural comparisons with other ADP-glucose pyrophosphorylases provided information to hypothesize that a Pro103-Arg115 loop is part of an activation path. It had strongly correlated movements with regions of the enzyme associated with regulation and ATP binding, and a network analysis showed that the optimal network pathways linking ATP and the activator binding Lys39 mainly involved residues of this loop. This hypothesis was biochemically tested by mutagenesis. We found that several alanine mutants of the Pro103-Arg115 loop had altered activation profiles for fructose-1,6-bisphosphate. Mutants P103A, Q106A, R107A, W113A, Y114A, and R115A had the most altered kinetic profiles, primarily characterized by a lack of response to fructose-1,6-bisphosphate. This loop is a distinct insertional element present only in allosterically regulated sugar nucleotide pyrophosphorylases that could have been acquired to build a triggering mechanism to link proto-allosteric and catalytic sites.
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1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Regulação Alostérica
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Escherichia coli
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Glucose-1-Fosfato Adenililtransferase
Idioma:
En
Revista:
Protein Sci
Assunto da revista:
BIOQUIMICA
Ano de publicação:
2015
Tipo de documento:
Article