Human UDP-galactose 4-epimerase. Accommodation of UDP-N-acetylglucosamine within the active site.
UDP-galactose 4-epimerase catalyzes the interconversion of UDP-galactose and UDP-glucose during normal galactose metabolism. One of the key structural features in the proposed reaction mechanism for the enzyme is the rotation of a 4'-ketopyranose intermediate within the active site pocket. Recently, the three-dimensional structure of the human enzyme with bound NADH and UDP-glucose was determined. Unlike that observed for the protein isolated from Escherichia coli, the human enzyme can also turn over UDP-GlcNAc to UDP-GalNAc and vice versa. Here we describe the three-dimensional structure of human epimerase complexed with NADH and UDP-GlcNAc. To accommodate the additional N-acetyl group at the C2 position of the sugar, the side chain of Asn-207 rotates toward the interior of the protein and interacts with Glu-199. Strikingly, in the human enzyme, the structural equivalent of Tyr-299 in the E. coli protein is replaced with a cysteine residue (Cys-307) and the active site volume for the human protein is calculated to be approximately 15% larger than that observed for the bacterial epimerase. This combination of a larger active site cavity and amino acid residue replacement most likely accounts for the inability of the E. coli enzyme to interconvert UDP-GlcNAc and UDP-GalNAc.
Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53705, USA.
J B
JB
Thoden
T M
TM
Wohlers
J L
JL
Fridovich-Keil
H M
HM
Holden
D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D008958:Models, Molecular; D009243:NAD / Q000378:metabolism; D011487:Protein Conformation; D014534:UDPglucose 4-Epimerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014531:Uridine Diphosphate Galactose / Q000378:metabolism; D014532:Uridine Diphosphate Glucose / Q000378:metabolism; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism*
D009243:NAD; D014531:Uridine Diphosphate Galactose; D014537:Uridine Diphosphate N-Acetylglucosamine; D014534:UDPglucose 4-Epimerase; D014532:Uridine Diphosphate Glucose