The Role of a Loop in the Non-catalytic Domain B on the Hydrolysis/Transglycosylation Specificity of the 4-& alpha;-Glucanotransferase from Thermotoga maritima


Por: Llopiz, Alexey, Ramirez-Martinez, Marco A., Olvera, Leticia, Xolalpa-Villanueva, Wendy, Pastor, Nina, Saab-Rincon, Gloria

Publicada: 1 oct 2023 Ahead of Print: 1 jul 2023
Resumen:
The mechanism by which glycoside hydrolases control the reaction specificity through hydrolysis or transglycosylation is a key element embedded in their chemical structures. The determinants of reaction specificity seem to be complex. We looked for structural differences in domain B between the 4-& alpha;-glucanotransferase from Thermotoga maritima (TmGTase) and the & alpha;-amylase from Thermotoga petrophila (TpAmylase) and found a longer loop in the former that extends towards the active site carrying a W residue at its tip. Based on these differences we constructed the variants W131G and the partial deletion of the loop at residues 120-124/128-131, which showed a 11.6 and 11.4-fold increased hydrolysis/transglycosylation (H/T) ratio relative to WT protein, respectively. These variants had a reduction in the maximum velocity of the transglycosylation reaction, while their affinity for maltose as the acceptor was not substantially affected. Molecular dynamics simulations allow us to rationalize the increase in H/T ratio in terms of the flexibility near the active site and the conformations of the catalytic acid residues and their associated pKas.

Filiaciones:
Llopiz, Alexey:
 Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Morelos, Cuernavaca, 62209, Mexico

Ramirez-Martinez, Marco A.:
 Centro de Investigación en Dinámica Celular, IICBA, Universidad Autónoma del Estado de Morelos, Morelos, Cuernavaca, 62209, Mexico

Olvera, Leticia:
 Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Morelos, Cuernavaca, 62209, Mexico

Xolalpa-Villanueva, Wendy:
 Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Morelos, Cuernavaca, 62209, Mexico

Pastor, Nina:
 Centro de Investigación en Dinámica Celular, IICBA, Universidad Autónoma del Estado de Morelos, Morelos, Cuernavaca, 62209, Mexico

Saab-Rincon, Gloria:
 Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Morelos, Cuernavaca, 62209, Mexico
ISSN: 15723887





PROTEIN JOURNAL
Editorial
Springer New York LLC, 233 SPRING ST, NEW YORK, NY 10013 USA, Estados Unidos America
Tipo de documento: Article
Volumen: 42 Número: 5
Páginas: 502-518
WOS Id: 001028766900001
ID de PubMed: 37464145
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