Isolation and characterization of the rml gene homologs from Porphyromonas gingivalis.
We cloned four genes from the Porphyromonas gingivalis chromosome, the gene products of which catalyze the anabolism of dTDP-L-rhamnose from D-glucose-1-phosphate when they were expressed in Escherichia coli. The amino acid sequences deduced from these genes showed significant homology to proteins encoded by the rml genes involved in dTDP-L-rhamnose biosynthesis in other gram-negative bacteria. Reverse transcriptase polymerase chain reaction analysis revealed that these four genes are expressed as a single transcript in P. gingivalis. To clarify the role of the rml gene homologs in this organism, construction of mutants defective in the rml gene homologs was attempted by allelic exchange. Unexpectedly, any mutants defective in the rml gene homologs were unable to be isolated, and the allelic exchange was possible only if the wild-type rml gene homologs were present on the chromosome. These results suggest that the rml gene homologs might be essential for the viability of P. gingivalis under the culture conditions used in this study.
Shibata
Y
Y
Department of Preventive Dentistry, Kyushu University Faculty of Dentistry, Fukuoka, Japan.
Yamashita
Y
Y
Nakano
Y
Y
Koga
T
T
D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification; D015139:Blotting, Southern; D002237:Carbohydrate Dehydrogenases / Q000235:genetics / Q000302:isolation & purification; D002238:Carbohydrate Epimerases / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D018274:Electroporation; D015870:Gene Expression; D005798:Genes, Bacterial; D006836:Hydro-Lyases / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D005810:Multigene Family; D009702:Nucleoside Diphosphate Sugars / Q000096:biosynthesis; D009713:Nucleotidyltransferases / Q000235:genetics / Q000302:isolation & purification; D016966:Porphyromonas gingivalis / Q000201:enzymology* / Q000235:genetics*; D015183:Restriction Mapping; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013942:Thymine Nucleotides / Q000096:biosynthesis
D001426:Bacterial Proteins; D009702:Nucleoside Diphosphate Sugars; D013942:Thymine Nucleotides; C073293:thymidine diphosphate rhamnose; D002237:Carbohydrate Dehydrogenases; C107144:dTDP-4-dehydrorhamnose reductase; D009713:Nucleotidyltransferases; C020587:glucose-1-phosphate thymidylyltransferase; D006836:Hydro-Lyases; C022771:dTDPglucose 4,6-dehydratase; D002238:Carbohydrate Epimerases; C020092:dTDP-4-ketorhamnose 3,5-epimerase