caa a22 4500 60551304X CHVBK 20210128100658.0 cr unu---uuuuu 210128e20150801xx s 000 0 eng 10.1007/s00894-015-2762-6 doi (NATIONALLICENCE)springer-10.1007/s00894-015-2762-6 Computational study enlightens the structural role of the alcohol acyltransferase DFGWG motif [Elektronische Daten] [Luis Morales-Quintana, María Moya-León, Raúl Herrera] Alcohol acyltransferases (AAT) catalyze the esterification reaction of alcohols and acyl-CoA into esters in fruits and flowers. Despite the high divergence between AAT enzymes, two important and conserved motifs are shared: the catalytic HxxxD motif, and the DFGWG motif. The latter is proposed to play a structural role; however, its function remains unclear. The DFGWG motif is located in loop 21 and stabilized by a hydrogen bond between residues Y52 and D381. Also, this motif is distant from the HxxxD motif, and most probably without a direct role in the substrate interaction. To evaluate the role of the DFGWG motif, in silico analysis was performed in the VpAAT1 protein. Three mutants (Y52F, D381A and D381E) were evaluated. Major changes (size and shape) in the solvent channels were found, although no differences were revealed in the entire 3D structure. Molecular dynamics simulations and docking studies described unfavorable energies for interaction of the mutant proteins with different substrates, as well as unfavored ligand orientations in the solvent channel. Additionally, we examined the contribution of different energetic parameters to the total free energy of protein-ligand complexes by the MM-GBSA method. The complexes differed mainly in their van der Waals contributions and have unfavorable electrostatic interactions. VpAAT1, Y52F and D381A mutants showed a dramatic reduction in the binding capacity to several substrates, which is related to differences in electrostatic potential on the protein surfaces, suggesting that D381 from the DFGWG motif and residue Y52 play a crucial role in maintenance of the adequate solvent channel structure required for catalysis. Graphical abstract Molecular docking, molecular dynamics (MD) simulations and MM-GBSA free energy calculations were employed to obtain quantitative estimates for the binding free energies of wild type Vasconcellea pubescens alcohol acyltransferase (VpAAT1-WT) and the protein mutants. Left VpAAT1 model structure in cartoon representation showing the solvent channel in the middle of the structure. Center, right Changes in shape and structure in the solvent channel of Y52F and D381A mutant proteins, respectively, compared to WT. The results obtained reveal that the interaction between D381 and Y52 residues is important for the maintenance of solvent channel structure Springer-Verlag Berlin Heidelberg, 2015 Alcohol acyltransferase nationallicence Ester biosynthesis nationallicence In silico site-directed mutagenesis nationallicence Molecular dynamics simulations nationallicence MM-GBSA nationallicence Vasconcellea pubescens nationallicence Morales-Quintana Luis Laboratorio de Fisiología Vegetal y Genética Molecular, Instituto de Ciencias Biológicas, Universidad de Talca, Casilla 747, Talca, Chile aut Moya-León María Laboratorio de Fisiología Vegetal y Genética Molecular, Instituto de Ciencias Biológicas, Universidad de Talca, Casilla 747, Talca, Chile aut Herrera Raúl Laboratorio de Fisiología Vegetal y Genética Molecular, Instituto de Ciencias Biológicas, Universidad de Talca, Casilla 747, Talca, Chile aut Journal of Molecular Modeling Springer Berlin Heidelberg 21/8(2015-08-01), 1-10 1610-2940 21:8<1 2015 21 894 https://doi.org/10.1007/s00894-015-2762-6 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00894-015-2762-6 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Morales-Quintana Luis Laboratorio de Fisiología Vegetal y Genética Molecular, Instituto de Ciencias Biológicas, Universidad de Talca, Casilla 747, Talca, Chile aut NATIONALLICENCE 700 1- Moya-León María Laboratorio de Fisiología Vegetal y Genética Molecular, Instituto de Ciencias Biológicas, Universidad de Talca, Casilla 747, Talca, Chile aut NATIONALLICENCE 700 1- Herrera Raúl Laboratorio de Fisiología Vegetal y Genética Molecular, Instituto de Ciencias Biológicas, Universidad de Talca, Casilla 747, Talca, Chile aut NATIONALLICENCE 773 0- Journal of Molecular Modeling Springer Berlin Heidelberg 21/8(2015-08-01), 1-10 1610-2940 21:8<1 2015 21 894