Kinetic analysis of the three-substrate reaction mechanism of an NRPS-independent siderophore (NIS) synthetase.
Methods Enzymol
; 702: 1-19, 2024.
Article
en En
| MEDLINE
| ID: mdl-39155107
ABSTRACT
The biosynthesis of many bacterial siderophores employs a member of a family of ligases that have been defined as NRPS-independent siderophore (NIS) synthetases. These NIS synthetases use a molecule of ATP to produce an amide linkage between a carboxylate and an amine. Commonly used carboxylate substrates include citrate or α-ketoglutarate, or derivatives thereof, while the amines are often hydroxamate derivatives of lysine or ornithine, or their decarboxylated forms cadaverine and putrescine. Enzymes that employ three substrates to catalyze a reaction may proceed through alternate mechanisms. Some enzymes use sequential mechanisms in which all three substrates bind prior to any chemical steps. In such mechanisms, substrates can bind in a random, ordered, or mixed fashion. Alternately, other enzymes employ a ping-pong mechanism in which a chemical step occurs prior to the binding of all three substrates. Here we describe an enzyme assay that will distinguish among these different mechanisms for the NIS synthetase, using IucA, an enzyme involved in the production of aerobactin, as the model system.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Péptido Sintasas
/
Sideróforos
Idioma:
En
Revista:
Methods Enzymol
Año:
2024
Tipo del documento:
Article
Pais de publicación:
Estados Unidos