Structural and functional analyses of the repressor, RbsR, of the ribose operon of Escherichia coli

Abstract

The DNA sequence encoding the rbs repressor protein, RbsR, has been determined. Amino acid sequence analyses of the product of an rbsR-lac2 fusion and of affinity-purified RbsR demonstrate that translation begins at an unusual codon, TTG, and that the initial amino acid is removed during maturation of the protein. DNA-binding assays indicate that RbsR binds to a region of perfect dyad symmetry spanning the rbs operon transcriptional start site and that the affinity for the rbs operator is reduced by addition of ribose, consistent with ribose being the inducer of the operon. RbsR is a member of a family of homologous repressor proteins having very similar DNA-binding sites and binding to similar operator sequences. [RbsR PIR accession number A41828.1 Keywords: Escherichia coli; repressors; ribose transport and utilization; transcription regulationThe rbs operon of Escherichia coli K12 encodes the highaffinity membrane transport system for ribose (Iida et al., 1984; Lopilato et al., 1984). Three open reading frames at the 5' end of the mRNA, designated rbsD, rbsA, and rbsC , are located in the region shown to be necessary for the membrane transport function (Iida et al., 1984;Lopilato et al., 1984). The coding region for the periplasmic ribose binding protein, rbsB, follows (Groarke et al., 1983), with that for ribokinase, rbsK , completing the set of genes that have been demonstrated to function as a single transcriptional unit (Lopilato et al., 1984).The gene encoding the repressor for the rbs operon, rbsR, was shown to be located downstream from rbsK, and evidence indicated that it is on a separate transcriptional unit from rbs (Lopilato et al ., 1984). During the sequence analysis, we found an open reading frame immediately downstream of rbsK that was clearly homologous to lacland galR . We have now completed the sequence analysis of this open reading frame and have shown that it encodes RbsR, a member of a family of repressor proteins. Functional studies have confirmed that the protein is expressed and binds to DNA sequences upstream of rbsD that conform to the consensus operator sequences for this family of repressors. These studies, together with the earlier genetic analyses (Lopilato et al., 1984), identify and locate the rbsR gene.

Carbohydrate metabolism in Bifidobacteria

“…Transcription of rbs is induced, thus facilitating ribose metabolism, in the presence of Dribose, which decreases the affinity of RbsR for its specific DNA operator in the promoter region of the rbs operon. This control mechanism is similar to RbsR regulation in E. coli [74]. In contrast, the binding activity RbsR from B. subtilis and Corynebacterium glutamicum does not appear to be affected by D-ribose [83,129].…”

Phylogenetic, structural and functional analyses of the LacI‐GalR family of bacterial transcription factors

“…2) were the only fully conserved residues in the multiple alignment of the 25 LGF proteins included in our study. Other residues in the H-T-H region ( The functional significance of some of these residues in specific members of the LGF is known and has been discussed [6,7,10,13,17]. For example, in PurR, S 14 (alignment position 22) hydrogen bonds to T17 (alignment position 25) accounting in part for the conservation of these residues.…”

“…Tile 3-dimensional structures of both the liganded and the unliganded forms of some of these proteins are known [6][7][8]19,20]. Outside of the regions of the LGF protein sequences depicted in Fig.…”

Structural homology between rbs repressor and ribose binding protein implies functional similarity

“…We show here that RbsR (Mauzy & Hermodson, 1992) is a member of a family of homologous repressor proteins, including Lad. The amino-terminal60 residues of each member of this protein family are predicted to form the bundle of four helices characteristic of proteins employing the helix-turn-helix DNA contact motif.…”

“…That issue is relevant for this family of repressor proteins, since LacI is known to form a tetramer when binding productively to the target DNA sequences (Platt et al, 1973). In addition, the symmetry of the operator sequences (Rolfes & Zalkin, 1990b;Mauzy & Hermodson, 1992) and almost universal precedent in other repressor systems makes it very likely that binding to the DNA requires at least a dimer of the repressor. The sites that bind inducer and those that are required to form dimers and tetramers in LacI all map to the region C-terminal to the 59-amino acid N-terminal headpiece (Platt et al, 1973), the region homologous to the binding proteins.…”