Scalable Chemoenzymatic Synthesis of Inositol Pyrophosphates.
Biochemistry
; 58(38): 3927-3932, 2019 09 24.
Article
em En
| MEDLINE
| ID: mdl-31461621
ABSTRACT
The inositol pyrophosphates (PP-InsPs) are an important group of cellular messengers that influence a broad range of biological processes. To elucidate the functions of these high-energy metabolites at the biochemical level, access to the purified molecules is required. Here, a robust and scalable strategy for the synthesis of various PP-InsPs [5PP-InsP5, 1PP-InsP5, and 1,5(PP)2-InsP4] is reported, relying on the highly active inositol hexakisphosphate kinase A from Entamoeba histolytica and the kinase domain of human diphosphoinositol pentakisphosphate kinase 2. A facile purification procedure using precipitation with Mg2+ ions and an optional strong anion exchange chromatography on an FPLC system afforded PP-InsPs in high purity. Furthermore, the newly developed protocol could be applied to simplify the synthesis of radiolabeled 5PP-InsP5-ß32P, which is a valuable tool for studying protein pyrophosphorylation. The chemoenzymatic method for obtaining PP-InsPs is readily amenable to both chemists and biologists and will thus foster future research on the multiple signaling functions of PP-InsP molecules.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Fosfotransferases (Aceptor do Grupo Fosfato)
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Difosfatos
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Fosfatos de Inositol
Idioma:
En
Ano de publicação:
2019
Tipo de documento:
Article