Crystal structures of Mycobacteria tuberculosis and Klebsiella pneumoniae UDP-galactopyranose mutase in the oxidised state and Klebsiella pneumoniae UDP-galactopyranose mutase in the (active) reduced state.
Uridine diphosphogalactofuranose (UDP-Galf) is the precursor of the d-galactofuranose sugar found in bacterial and parasitic cell walls, including those of many pathogens. UDP-Galf is made from UDP-galactopyranose by the enzyme UDP-galactopyranose mutase. The enzyme requires the reduced FADH- co-factor for activity. The structure of the Mycobacterium tuberculosis mutase with FAD has been determined to 2.25 A. The structures of Klebsiella pneumoniae mutase with FAD and with FADH- bound have been determined to 2.2 A and 2.35 A resolution, respectively. This is the first report of the FADH(-)-containing structure. Two flavin-dependent mechanisms for the enzyme have been proposed, one, which involves a covalent adduct being formed at the flavin and the other based on electron transfer. Using our structural data, we have examined the two mechanisms. The electron transfer mechanism is consistent with the structural data, not surprisingly, since it makes fewer demands on the precise positioning of atoms. A model based on a covalent adduct FAD requires repositioning of the enzyme active site and would appear to require the isoalloxazine ring of FADH- to buckle in a particular way. However, the FADH- structure reveals that the isoalloxazine ring buckles in the opposite sense, this apparently requires the covalent adduct to trigger profound conformational changes in the protein or to buckle the FADH- opposite to that seen in the apo structure.
Beis
Konstantinos
K
Centre for Biomolecular Sciences, University of St Andrews, North Haugh, St Andrews, Fife, Scotland KY16 9ST, UK.
Srikannathasan
Velupillai
V
Liu
Huanting
H
Fullerton
Stephen W B
SW
Bamford
Vicki A
VA
Sanders
David A R
DA
Whitfield
Chris
C
McNeil
Mike R
MR
Naismith
James H
JH
D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D005415:Flavins / Q000494:pharmacology; D019751:Intramolecular Transferases / Q000737:chemistry* / Q000378:metabolism*; D007711:Klebsiella pneumoniae / Q000201:enzymology*; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D010084:Oxidation-Reduction; D020836:Protein Structure, Quaternary; D016415:Sequence Alignment
D005415:Flavins; D008024:Ligands; D019751:Intramolecular Transferases; C098051:UDP-galactopyranose mutase