PMID:15598657
| Citation |
Zhang, Z, Tsujimura, M, Akutsu, J, Sasaki, M, Tajima, H and Kawarabayasi, Y (2005) Identification of an extremely thermostable enzyme with dual sugar-1-phosphate nucleotidylyltransferase activities from an acidothermophilic archaeon, Sulfolobus tokodaii strain 7. J. Biol. Chem. 280:9698-705 |
|---|---|
| Abstract |
L-rhamnose is an essential component of the cell wall and plays roles in mediating virulence and adhesion to host tissues in many microorganisms. Glucose-1-phosphate thymidylyltransferase (RmlA, EC 2.7.7.24) catalyzes the first reaction of the four-step pathway of L-rhamnose biosynthesis, producing dTDP-D-glucose from dTTP and glucose-1-phosphate. Three RmlA homologues of varying size have been identified in the genome of a thermophilic archaeon, Sulfolobus tokodaii strain 7. In this study, we report the heterologous expression of the largest homologue (a 401 residue-long ST0452 protein) and characterization of its thermostable activity. RmlA enzymatic activity of this protein was detected from 65 to 100 degrees C, with a half-life of 60 min at 95 degrees C and 180 min at 80 degrees C. Analysis of a deletion mutant lacking the 170-residue C-terminal domain indicated that this region has an important role in the thermostability and activity of the protein. Analyses of substrate specificity indicated that the enzymatic activity of the full-length protein is capable of utilizing alpha-D-glucose-1-phosphate and N-acetyl-D-glucosamine-1-phosphate but not alpha-D-glucosamine-1-phosphate. However, the protein is capable of utilizing all four deoxyribonucleoside triphosphates and UTP. Thus, the ST0452 protein is an enzyme containing both glucose-1-phosphate thymidylyltransferase and N-acetyl-D-glucosamine-1-phosphate uridylyltransferase activities. This is the first report of a thermostable enzyme with dual sugar-1-phosphate nucleotidylyltransferase activities. |
| Links |
PubMed Online version:10.1074/jbc.M411211200 |
| Keywords |
Amino Acid Sequence; Archaeal Proteins/chemistry; Archaeal Proteins/genetics; Archaeal Proteins/metabolism; Base Sequence; Cloning, Molecular; Conserved Sequence; DNA Primers; Enzyme Stability; Hot Temperature; Molecular Sequence Data; Nucleotidyltransferases/chemistry; Nucleotidyltransferases/genetics; Nucleotidyltransferases/metabolism; Open Reading Frames; Recombinant Proteins/chemistry; Recombinant Proteins/metabolism; Sequence Alignment; Sequence Deletion; Sequence Homology, Amino Acid; Sulfolobus/enzymology; Thermodynamics |
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