Establishing the Catalytic Mechanism of Human Pancreatic alpha-Amylase with QM/MM Methods

• Tipo: Artigo em Revista Científica Internacional
• Data: 2015

Autores

Gaspar P Pinto (Autor) Outra; Natercia F Bras (Autor) Outra; Marta A S Perez (Autor) Outra; Pedro A Fernandes (Autor) FCUP; Nino Russo (Autor) Outra; Maria J Ramos (Autor) FCUP; Marirosa Toscano (Autor) Outra

Revista

Título: Journal of Chemical Theory and Computation

Vol. 11

Páginas: 2508-2516

ISSN: 1549-9618

Editora: American Chemical Society

Indexação

ISI Web of Knowledge - 29 Citações

Scopus - 28 Citações

Classificação Científica

FOS: Ciências exactas e naturais > Química

Outras Informações

ID Authenticus: P-00G-BRW

DOI: 10.1021/acs.jctc.5b00222

Abstract (EN): In this work, we studied the catalytic mechanism of human pancreatic alpha-amylase (HPA). Our goal was to determine the catalytic mechanism of HPA with atomic detail using computational methods. We demonstrated that the HPA catalytic mechanism consists of two steps, the first of which (glycosylation step) involves breaking the glycosidic bond to culminate in the formation of a covalent intermediate. The second (deglycosylation step) :consists of the addition of a water molecule to release the enzyme/substrate covalent intermediate, completing the hydrolysis of the sugar. The active site was very open to the solvent. Our mechanism basically differs from the previously proposed mechanism by having two water molecules instead of only one near the active site that participate in the mechanism. We also demonstrate the relevant role of the three catalytic amino acids, two aspartate residues and a glutamate (D197, E233, and D300), during catalysis. It was also Shown that the rate limiting step was glycosylation, and its activation energy was in agreement with experimental values obtained for HPA. The experimental activation energy was 14.4 kcal mol(-1), and the activation energy obtained computationally was 15.1 kcal mol(-1).

Idioma: Inglês

Tipo (Avaliação Docente): Científica

Nº de páginas: 9