Acetylcholinesterases from Leaf-Cutting ant Atta sexdens: Purification, Characterization, and Capillary Reactors for On-Flow Assays.
Enzyme Res
; 2019: 6139863, 2019.
Article
em En
| MEDLINE
| ID: mdl-31354985
Acetylcholinesterase (AChE) is responsible for catalyzing the hydrolysis of the neurotransmitter acetylcholine (ACh) leading to acetate and choline (Ch) release. The inhibition of AChE produces a generalized synaptic collapse that can lead to insect death. Herein we report for the first time the isolation of two AChEs from Atta sexdens which were purified by sulphate ammonium precipitation followed by ion exchange chromatography. AsAChE-A and AsAChE-B enzymes have optimum pH of 9.5 and 9.0 and higher activities in 30/50°C and 20°C, respectively, using acetylthiocholine (ATCh) as substrate. Immobilized capillary enzyme reactors (ICERs) were obtained for both enzymes (AsAChE-A-ICER and AsAChE-B-ICER) and their activities were measured by LC-MS/MS through hydrolysis product quantification of the natural substrate ACh. The comparison of activities by LC-MS/MS of both AChEs using ACh as substrate showed that AsAChE-B (free or immobilized) had the highest affinity. The inverse result was observed when the colorimetric assay (Elman method) was used for ATCh as substrate. Moreover, by mass spectrometry and phylogenetic studies, AsAChE-A and AsAChE-B were classified as belonging to AChE-2 and AChE-1 classes, respectively.
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1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Enzyme Res
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
Brasil
País de publicação:
Estados Unidos