The gene encoding the alternative thymidylate synthase ThyX is regulated by sigma factor SigB in Corynebacterium glutamicum ATCC 13032.
Both ThyA and ThyX proteins catalyze the transfer of the methyl group from methylenetetrahydrofolate (CH(2) H(4) -folate) to dUMP, forming dTMP. To estimate the relative steady state expression levels of ThyA and ThyX, Western blot analysis was performed using ThyA or ThyX antiserum on total protein from the wild-type, ΔthyX, and thyX-complemented strains of Corynebacterium glutamicum. The level of ThyA decreased gradually during the stationary growth phase but that of ThyX was maintained steadily. Whereas the expression level of ThyA in a ΔsigB strain was comparable to that of the wild-type, the level of ThyX was significantly diminished in the deletion mutant and was restored to that of the wild-type in the complemented strain, indicating that the level of ThyX was regulated by SigB. Growth of the C. glutamicum ΔsigB strain was dependent upon coupling activity of dihydrofolate reductase (DHFR) with ThyA for the synthesis of thymidine, and thus showed sensitivity to the inhibition of DHFR by the experimental inhibitor, WR99210-HCl. These results suggested that the relative levels of ThyA and ThyX differ in response to different growth phases and that SigB is necessary for maintenance of the level of ThyX during transition into the stationary growth phase.
Department of Biology, Kyung Hee University, Seoul, Korea.
Sukhyeong
S
Cho
Seokbin
S
Yang
Hogun
H
Rhie
D001426:Bacterial Proteins / Q000378:metabolism*; D015153:Blotting, Western; D048230:Corynebacterium glutamicum / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D003470:Culture Media / Q000378:metabolism; D057075:Enzyme Assays; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D005798:Genes, Bacterial; D017930:Genes, Reporter; D008826:Microbial Sensitivity Tests; D016296:Mutagenesis; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012808:Sigma Factor / Q000378:metabolism*; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D013940:Thymidylate Synthase / Q000235:genetics / Q000378:metabolism*; D014227:Triazines / Q000494:pharmacology; D001616:beta-Galactosidase / Q000378:metabolism
D001426:Bacterial Proteins; D003470:Culture Media; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; C079011:SigB protein, Bacteria; D012808:Sigma Factor; D014227:Triazines; C006201:BRL 6231; D013762:Tetrahydrofolate Dehydrogenase; D013940:Thymidylate Synthase; D001616:beta-Galactosidase; D013936:Thymidine