The crystal structure of dTDP-D-Glucose 4,6-dehydratase (RmlB) from Salmonella enterica serovar Typhimurium, the second enzyme in the dTDP-l-rhamnose pathway.
l-Rhamnose is a 6-deoxyhexose that is found in a variety of different glycoconjugates in the cell walls of pathogenic bacteria. The precursor of l-rhamnose is dTDP-l-rhamnose, which is synthesised from glucose- 1-phosphate and deoxythymidine triphosphate (dTTP) via a pathway requiring four enzymes. Significantly this pathway does not exist in humans and all four enzymes therefore represent potential therapeutic targets. dTDP-D-glucose 4,6-dehydratase (RmlB; EC 4.2.1.46) is the second enzyme in the dTDP-L-rhamnose biosynthetic pathway. The structure of Salmonella enterica serovar Typhimurium RmlB had been determined to 2.47 A resolution with its cofactor NAD(+) bound. The structure has been refined to a crystallographic R-factor of 20.4 % and an R-free value of 24.9 % with good stereochemistry.RmlB functions as a homodimer with monomer association occurring principally through hydrophobic interactions via a four-helix bundle. Each monomer exhibits an alpha/beta structure that can be divided into two domains. The larger N-terminal domain binds the nucleotide cofactor NAD(+) and consists of a seven-stranded beta-sheet surrounded by alpha-helices. The smaller C-terminal domain is responsible for binding the sugar substrate dTDP-d-glucose and contains four beta-strands and six alpha-helices. The two domains meet to form a cavity in the enzyme. The highly conserved active site Tyr(167)XXXLys(171) catalytic couple and the GlyXGlyXXGly motif at the N terminus characterise RmlB as a member of the short-chain dehydrogenase/reductase extended family. The quaternary structure of RmlB and its similarity to a number of other closely related short-chain dehydrogenase/reductase enzymes have enabled us to propose a mechanism of catalysis for this important enzyme.
Centre for Biomolecular Sciences, The University, North Haugh, St Andrews Fife, KY16 9ST, Scotland.
S T
ST
Allard
M F
MF
Giraud
C
C
Whitfield
M
M
Graninger
P
P
Messner
J H
JH
Naismith
D000595:Amino Acid Sequence; D001665:Binding Sites; D002384:Catalysis; D018360:Crystallography, X-Ray; D006836:Hydro-Lyases / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D009243:NAD / Q000737:chemistry*; D009702:Nucleoside Diphosphate Sugars / Q000378:metabolism; D009711:Nucleotides / Q000737:chemistry; D011487:Protein Conformation; D019779:Salmonella enterica / Q000737:chemistry* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012703:Serotyping; D013942:Thymine Nucleotides / Q000378:metabolism
D009702:Nucleoside Diphosphate Sugars; D009711:Nucleotides; D013942:Thymine Nucleotides; D009243:NAD; C073293:thymidine diphosphate rhamnose; D006836:Hydro-Lyases; C022771:dTDPglucose 4,6-dehydratase