Analysis of the reaction coordinate of alpha-L-fucosidases: a combined structural and quantum mechanical approach.
The enzymatic hydrolysis of alpha-L-fucosides is of importance in cancer, bacterial infections, and fucosidosis, a neurodegenerative lysosomal storage disorder. Here we show a series of snapshots along the reaction coordinate of a glycoside hydrolase family GH29 alpha-L-fucosidase unveiling a Michaelis (ES) complex in a (1)C(4) (chair) conformation and a covalent glycosyl-enzyme intermediate in (3)S(1) (skew-boat). First principles metadynamics simulations on isolated alpha-L-fucose strongly support a (1)C(4)<-->(3)H(4)<-->(3)S(1) conformational itinerary for the glycosylation step of the reaction mechanism and indicate a strong "preactivation" of the (1)C(4) complex to nucleophilic attack as reflected by free energy, C1-O1/O5-C1 bond length elongation/reduction, C1-O1 bond orientation, and positive charge development around the anomeric carbon. Analysis of an imino sugar inhibitor is consistent with tight binding of a chair-conformed charged species.
Lammerts van Bueren
Alicia
A
York Structural Biology Laboratory, Department of Chemistry, The University of York, York YO10 5YW, UK.
Ardèvol
Albert
A
Fayers-Kerr
Jennifer
J
Luo
Bo
B
Zhang
Yongmin
Y
Sollogoub
Matthieu
M
Blériot
Yves
Y
Rovira
Carme
C
Davies
Gideon J
GJ
D001439:Bacteroides / Q000201:enzymology; D005643:Fucose / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D011487:Protein Conformation; D011789:Quantum Theory; D017386:Sequence Homology, Amino Acid; D013816:Thermodynamics; D005644:alpha-L-Fucosidase / Q000737:chemistry* / Q000378:metabolism*
D005643:Fucose; D005644:alpha-L-Fucosidase