Some features of DNA-binding proteins involved in the regulation of the Streptomyces aureofaciens gap gene, encoding glyceraldehyde-3-phosphate dehydrogenase.
A gapR gene, encoding a protein similar to the AraC/XylS family of bacterial transcriptional regulators, was previously identified upstream of the gap gene, coding for glyceraldehyde-3-phosphate dehydrogenase in Streptomyces aureofaciens. The GapR protein overproduced in Escherichia coli was shown to bind to the gap-P promoter region. Using the gel mobility shift assay with cell-free protein extracts from different developmental stages of S. aureofaciens, we identified several other proteins, in addition to GapR, that specifically bound to the S. aureofaciens gap-P promoter region. When cell-free extracts from S. aureofaciens cultivated in liquid medium with glucose were analyzed, only one complex corresponding to GapR was detected. A new protein interacting with the gap-P promoter was detected in stationary culture of S. aureofaciens grown in the presence of mannitol as carbon sources. The GapR protein was partially purified from S. aureofaciens cultivated in liquid medium containing glucose and used for binding studies. DNA footprinting analysis revealed an identical protected region as previously identified for the GapR protein overproduced from Escherichia coli. The direct role of the GapR protein in the regulation of gap expression in S. aureofaciens in vivo was confirmed but regulation of gap expression seems to be more complex, possibly involving other regulatory protein(s), depending on the developmental stage of S. aureofaciens.
Homerová
D
D
Institute of Molecular Biology, Slovak Academy of Sciences, 842 51 Bratislava, Slovakia.
Sprusanský
O
O
Kutejová
E
E
Kormanec
J
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D005798:Genes, Bacterial; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015183:Restriction Mapping; D013304:Streptomyces aureofaciens / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; C435219:GapR protein, Streptomyces aureofaciens; D011994:Recombinant Proteins; D015534:Trans-Activators; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases