Your browser doesn't support javascript.
loading
Kinetic characterization and phylogenetic analysis of human ADP-dependent glucokinase reveal new insights into its regulatory properties.
Herrera-Morandé, Alejandra; Vallejos-Baccelliere, Gabriel; Cea, Pablo A; Zamora, Ricardo A; Cid, Dixon; Maturana, Pablo; González-Ordenes, Felipe; Castro-Fernández, Víctor; Guixé, Victoria.
Afiliação
  • Herrera-Morandé A; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile. Electronic address: alejandra.herrera@uautonoma.cl.
  • Vallejos-Baccelliere G; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile. Electronic address: gvallejos@ug.uchile.cl.
  • Cea PA; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile.
  • Zamora RA; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile.
  • Cid D; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile.
  • Maturana P; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile.
  • González-Ordenes F; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile.
  • Castro-Fernández V; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile.
  • Guixé V; Laboratorio de Bioquímica y Biología Molecular, Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago, Chile. Electronic address: vguixe@uchile.cl.
Arch Biochem Biophys ; 741: 109602, 2023 06.
Article em En | MEDLINE | ID: mdl-37084804
ABSTRACT
Although ADP-dependent sugar kinases were first described in archaea, at present, the presence of an ADP-dependent glucokinase (ADP-GK) in mammals is well documented. This enzyme is mainly expressed in hematopoietic lineages and tumor tissues, although its role has remained elusive. Here, we report a detailed kinetic characterization of the human ADP-dependent glucokinase (hADP-GK), addressing the influence of a putative signal peptide for endoplasmic reticulum (ER) destination by characterizing a truncated form. The truncated form revealed no significant impact on the kinetic parameters, showing only a slight increase in the Vmax value, higher metal promiscuity, and the same nucleotide specificity as the full-length enzyme. hADP-GK presents an ordered sequential kinetic mechanism in which MgADP is the first substrate to bind and AMP is the last product released, being the same mechanism described for archaeal ADP-dependent sugar kinases, in agreement with the protein topology. Substrate inhibition by glucose was observed due to sugar binding to nonproductive species. Although Mg2+ is an essential component for kinase activity, it also behaves as a partial mixed-type inhibitor for hADP-GK, mainly by decreasing the MgADP affinity. Regarding its distribution, phylogenetic analysis shows that ADP-GK's are present in a wide diversity of eukaryotic organisms although it is not ubiquitous. Eukaryotic ADP-GKs sequences cluster into two main groups, showing differences in the highly conserved sugar-binding motif reported for archaeal enzymes [NX(N)XD] where a cysteine residue is found instead of asparagine in a significant number of enzymes. Site directed mutagenesis of the cysteine residue by asparagine produces a 6-fold decrease in Vmax, suggesting a role for this residue in the catalytic process, probably by facilitating the proper orientation of the substrate to be phosphorylated.
Assuntos
Palavras-chave

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Asparagina / Cisteína Limite: Humans Idioma: En Revista: Arch Biochem Biophys Ano de publicação: 2023 Tipo de documento: Article

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Asparagina / Cisteína Limite: Humans Idioma: En Revista: Arch Biochem Biophys Ano de publicação: 2023 Tipo de documento: Article