Structural analysis of Thermus thermophilus HB27 mannosyl-3-phosphoglycerate synthase provides evidence for a second catalytic metal ion and new insight into the retaining mechanism of glycosyltransferases.
Mannosyl-3-phosphoglycerate synthase is a glycosyltransferase involved in the two-step synthetic pathway of mannosylglycerate, a compatible solute that accumulates in response to salt and/or heat stresses in many microorganisms thriving in hot environments. The three-dimensional structure of mannosyl-3-phosphoglycerate synthase from Thermus thermophilus HB27 in its binary complex form, with GDP-alpha-D-mannose and Mg(2+), shows a second metal binding site, about 6 A away from the mannose moiety. Kinetic and mutagenesis studies have shown that this metal site plays a role in catalysis. Additionally, Asp(167) in the DXD motif is found within van der Waals contact distance of the C1' atom in the mannopyranose ring, suggesting its action as a catalytic nucleophile, either in the formation of a glycosyl-enzyme intermediate according to the double-displacement S(N)2 reaction mechanism or in the stabilization of the oxocarbenium ion-like intermediate according to the D(N)*A(Nss) (S(N)i-like) reaction mechanism. We propose that either mechanism may occur in retaining glycosyltransferases with a GT-A fold, and, based on the gathered structural information, we identified an extended structural signature toward a common scaffold between the inverting and retaining glycosyltransferases.
Gonçalves
Susana
S
Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Apartado 127, 2781-901 Oeiras, Portugal.
Borges
Nuno
N
Esteves
Ana M
AM
Victor
Bruno L
BL
Soares
Cláudio M
CM
Santos
Helena
H
Matias
Pedro M
PM
D002384:Catalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray / Q000379:methods; D016695:Glycosyltransferases / Q000737:chemistry*; D007477:Ions; D007700:Kinetics; D008358:Mannose / Q000737:chemistry; D008364:Mannosyltransferases / Q000737:chemistry*; D008670:Metals / Q000737:chemistry*; D008954:Models, Biological; D008956:Models, Chemical; D008958:Models, Molecular; D008968:Molecular Conformation; D016296:Mutagenesis; D016963:Thermus thermophilus / Q000378:metabolism*
D007477:Ions; D008670:Metals; D016695:Glycosyltransferases; D008364:Mannosyltransferases; C441639:mannosyl-3-phosphoglycerate synthase; D008358:Mannose