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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The biosynthesis of methionine in Escherichia coli is under complex regulation. The repression of the biosynthetic pathway by methionine is mediated by a repressor protein (MetJ protein) and S-adenosyl-methionine which functions as a corepressor for the MetJ protein. Recently, a new regulatory locus, metR, has been identified. The MetR protein is required for both metE and metH gene expression, and functions as a transactivator of transcription of these genes. MetR is a unique prokaryotic transcription activator in that it possesses a leucine zipper motif, first described for eukaryotic DNA-binding proteins. The transcriptional activity of MetR is modulated by homocysteine, the metabolic precursor of methionine. Finally, it is known that vitamin B12 can repress expression of the metE gene. This effect is mediated by the MetH holoenzyme, which contains a cobamide prosthetic group.
Mol Microbiol 1991 Jul
PMID:Regulation of methionine synthesis in Escherichia coli. 194 95

We have altered the amino acid sequence of the lac repressor one residue at a time by utilizing a collection of nonsense suppressors that permit the insertion of 13 different amino acids in response to the amber (UAG) codon, as well as an additional amino acid in response to the UGA codon. We used this collection to suppress nonsense mutations at 141 positions in the lacI gene, which encodes the 360 amino acid long lac repressor, including 53 new nonsense mutations which we constructed by oligonucleotide-directed mutagenesis. This method has generated over 1600 single amino acid substitutions in the lac repressor. We have cataloged the effects of these replacements and have interpreted the results with the objective of gaining a better understanding of lac repressor structure, and protein structure in general. The DNA binding domain of the repressor, involving the amino-terminal 59 amino acids, is extremely sensitive to substitution, with 70% of the replacements resulting in the I- phenotype. However, the remaining 301 amino acid core of the repressor is strikingly tolerant of substitutions, with only 30% of the amino acids introduced causing the I- phenotype. This analysis reveals the location of sites in the protein involved in inducer binding, tighter binding to operator and thermal stability, and permits a virtual genetic image reconstruction of the lac repressor protein.
J Mol Biol 1990 Mar 20
PMID:Genetic studies of the lac repressor. XIII. Extensive amino acid replacements generated by the use of natural and synthetic nonsense suppressors. 215 24

The Escherichia coli cytR-encoded repressor protein (CytR) controls the expression of several genes involved in nucleoside and deoxynucleoside uptake and metabolism. The cytR promoter was identified by determining the transcriptional initiation site of the cytR gene. A chromosomal cytR-lacZ+ operon fusion was isolated and used to study the regulation of cytR. We show that cytR expression is negatively controlled by the CytR protein and positively affected by the cAMP/CAP complex. Footprinting studies with purified CAP protein revealed two CAP binding sites upstream of the cytR promoter. A previously described mutation (cytR*) in the cloned cytR gene, which results in the phenotypic suppression of a CytR operator mutation in the tsx P2 promoter, was analysed. DNA sequence analysis of the cytR* mutation revealed a G-C to an A-T base pair transition at position -34 bp relative to the translational initiation site of cytR. This point mutation activates a cryptic promoter that is stronger than the wild-type cytR promoter and leads to overproduction of the CytR repressor.
Mol Microbiol 1990 Mar
PMID:Transcriptional regulation of the cytR repressor gene of Escherichia coli: autoregulation and positive control by the cAMP/CAP complex. 216 67

The biochemical properties of the recA430 protein have been examined and compared to those of wild-type recA protein. We find that, while the recA430 protein possesses ssDNA-dependent rATP activity, this activity is inhibited by the Escherichia coli single-stranded DNA binding protein (SSB protein) under many conditions that enhance wild-type recA protein rATPase hydrolysis. Stimulation of rATPase activity by SSB protein is observed only at high concentrations of both rATP (greater than 1 mM) and recA430 protein (greater than 5 microM). In contrast, stimulation of ssDNA-dependent dATPase activity by SSB protein is less sensitive to protein and nucleotide concentration. Consistent with the nucleotide hydrolysis data, recA430 protein can carry out DNA strand exchange in the presence of either rATP or dATP. However, in the presence of rATP, both the rate and the extent of DNA strand exchange by recA430 protein are greatly reduced compared to wild-type recA protein and are sensitive to recA430 protein concentration. This reduction is presumably due to the inability of recA430 protein to compete with SSB protein for ssDNA binding sites under these conditions. The cleavage of lexA repressor protein by recA430 protein is also sensitive to the nucleotide cofactor present and is completely inhibited by SSB protein when rATP is the cofactor but not when dATP is used. Finally, the steady-state affinity and the rate of association of the recA430 protein-ssDNA complex are reduced, suggesting that the mutation affects the interaction of the ATP-bound form of recA protein with ssDNA. This alteration is the likely molecular defect responsible for inhibition of recA430 protein rATP-dependent function by SSB protein. The biochemical properties observed in the presence of dATP and SSB protein, i.e. the reduced levels of both DNA strand exchange activity and cleavage of lexA repressor protein, are consistent with the phenotypic behavior of recA430 mutations.
J Mol Biol 1990 Feb 20
PMID:Biochemical properties of the Escherichia coli recA430 protein. Analysis of a mutation that affects the interaction of the ATP-recA protein complex with single-stranded DNA. 217 66

The osteolytic toxin of Pasteurella multocida induces bone resorption in vivo and in vitro (Foged et al., 1988; Kimman et al., 1987). In this report the toxin-encoding toxA gene is sequenced, and the deduced primary structure of the toxin shows a protein of 1285 amino acids, containing a striking homology to a metal-binding motif. Evidence that expression of the toxA gene is repressed at a transcriptional level in Escherichia coli is presented. Repression could be abolished either by deletion of a region upstream of toxA, or by a putative frame-shift mutation in the same region. The repressor protein encoded within this region was efficient in trans, and was named TxaR.
Mol Microbiol 1990 May
PMID:The complete nucleotide sequence of the Pasteurella multocida toxin gene and evidence for a transcriptional repressor, TxaR. 220 70

The yeast GCN4 transcriptional activator protein binds as a dimer to a dyad-symmetric sequence, indicative of a protein-DNA complex in which two protein monomers interact with adjacent half-sites. However, the optimal GCN4 recognition site, ATGA(C/G)TCAT, is inherently asymmetric because it contains an odd number of base pairs and because mutation of the central C.G base pair strongly reduces specific DNA binding. From this asymmetry, we suggested previously that GCN4 interacts with nonequivalent and possibly overlapping half-sites (ATGAC and ATGAG) that have different affinities. Here, we examine the nature of GCN4 half-sites by creating symmetrical derivatives of the optimal GCN4 binding sequence that delete or insert a single base pair at the center of the site. In vitro, GCN4 bound efficiently to the sequence ATGACGTCAT, whereas it failed to bind to ATGAGCTCAT or ATGATCAT. These observations strongly suggest that (i) GCN4 specifically recognizes the central base pair, (ii) the optimal half-site for GCN4 binding is ATGAC, not ATGAG, and (iii) GCN4 is a surprisingly flexible protein that can accommodate the insertion of a single base pair in the center of its compact binding site. The ATGACGTCAT sequence strongly resembles sites bound by the yeast and mammalian ATF/CREB family of proteins, suggesting that GCN4 and the ATF/CREB proteins recognize similar half-sites but have different spacing requirements. Unexpectedly, in the context of the his3 promoter, the ATGACGTCAT derivative reduced transcription below the basal level in a GCN4-independent manner, presumably reflecting DNA binding by a distinct ATF/CREB-like repressor protein. In other promoter contexts, however, the same site acted as a weak upstream activating sequence.
Mol Cell Biol 1990 Oct
PMID:Mutations that define the optimal half-site for binding yeast GCN4 activator protein and identify an ATF/CREB-like repressor that recognizes similar DNA sites. 220 5

The half-life of ribosomal protein operon L11 mRNA in vivo was measured during exponential growth by following the kinetics of incorporation of radioactive precursors into L11 mRNA transcribed from multi-copy plasmids. The degree of translational feedback regulation by L1, the L11 operon-specific translational repressor protein, was changed by altering the site on the "L11 mRNA" where L1 interacts. The half-life of the overproduced L11 mRNA increased by about fivefold when translational repression was abolished, while the half-life of mRNA from the spc ribosomal protein operon, which is not translationally regulated by L1, stayed constant. Furthermore, the half-life of L11 operon mRNA carrying an additional mutation in the ribosome binding site that abolishes translation remains short. This indicates that the change in half-life observed during increased gene dosage is due to translational repression by L1 and is probably a consequence of L1 blocking translation of L11 mRNA and not due to some nucleolytic activity mediated by L1.
J Mol Biol 1986 Apr 05
PMID:Changes in the half-life of ribosomal protein messenger RNA caused by translational repression. 242 54

The regulation of expression of the Tn1721-encoded tetracycline-resistance determinant is described at the molecular level. The transcriptional control element consists of overlapping divergent promoters, which are negatively regulated by two operators with nearly identical sequence. The mRNA for the regulatory gene tetR is translated without a ribosome-binding site. This result is confirmed by S1 nuclease mapping and RNA sequencing of the tetR mRNA. The start nucleotide for transcription of this mRNA is the adenosine residue of the sequence 5'-AUG. Determination of the N-terminal amino acid sequence of the purified Tet repressor proves that this AUG is the initiation codon for translation. The Tet repressor protein is further used to map the two tet operators by DNase I footprinting. Tight contacts of the protein to the N-7 positions of two guanosine residues in each operator are determined from methylation protection experiments with dimethylsulfate. The differential regulation and positive control of transcription of the tetR gene that is possible with this arrangement of promoters and operators is discussed.
J Mol Biol 1986 Jun 20
PMID:Expression, purification and operator binding of the transposon Tn1721-encoded Tet repressor. 243 Nov 53

A quantitative model for the regulation of replication of the low copy number IncFII plasmid NR1 in the Escherichia coli cell division cycle has been developed. The initiation of NR1 replication requires a cis-acting initiator protein whose synthesis is regulated by several mechanisms. The NR1 regulatory processes include co-operative protein-protein interactions in the formation of an active transcription repressor, the interaction of repressor with a rightward operator site in the control of transcription of the initiator gene, and the interaction of an inhibitor RNA transcript with the initiator mRNA in the control of translation of the initiation protein. A statistical thermodynamic model was used to predict probable configurations of the regulatory processes in a single growing cell. These probabilities were coupled by a kinetic model to the events of the cell cycle, such as initiation of mRNA transcription and protein translation, and the initiation of plasmid DNA replication. Parameter values were chosen so that the simulated values for plasmid copy number and the intracellular concentrations of repressor protein and mRNA agreed with experimentally determined estimates. A number of different copy number mutants that have altered one or another of the regulatory processes were simulated by the model. The contributions of each of the regulatory processes toward the overall stability of inheritance of plasmid NR1 in a population of cells in culture were examined. These simulations predict a very stable pattern of inheritance for plasmid NR1 despite its low copy number, in agreement with experimental observation.
J Mol Biol 1986 Dec 05
PMID:Regulation of IncFII plasmid DNA replication. A quantitative model for control of plasmid NR1 replication in the bacterial cell division cycle. 243 19

The spc ribosomal protein operon of Escherichia coli is feedback-regulated by ribosomal protein S8, a translational repressor. We have analyzed the region of the spc mRNA that is responsible for this regulation. First, we have established that the S8 target site on the mRNA is near the translation start site of the third gene encoding ribosomal protein L5 in the operon. This was done by constructing hybrid plasmids carrying spc operon ribosomal protein genes under lac transcriptional control, as well as their deletion derivatives, and carrying out both in vivo and in vitro protein synthesis experiments. Next, the secondary structure of this region was studied by analyzing 5' end-labeled RNA synthesized from the phage SP6 promoter using structure-specific nucleases. A secondary structure model consistent with the results was deduced with the aid of a computer prediction of RNA folding. In addition, we cloned and sequenced the corresponding region from Salmonella typhimurium, Proteus vulgaris and Serratia marcescens and found five "compensating" substitutions that support some of the deduced helical structures of mRNA. None of the base changes was inconsistent with the deduced secondary structure model. Finally, site-directed mutagenesis experiments have identified bases important for regulation, including two base-paired sites representing each of two helical regions. This has led to the conclusion that some specific nucleotide residues located between these two helical regions are directly involved in S8 recognition, and that the function of the two helical regions is to maintain the proper orientation of these nucleotide residues. Comparison of the structure of the S8 target site on the spc mRNA with the known S8 binding site on rRNA has revealed a striking similarity in both primary and secondary structures. In particular, primary sequences of rRNA conserved among distantly related bacterial species in this region is found to be identical with the sequences at the corresponding positions in mRNA. These results suggest that the same structural features of the S8 repressor protein are involved in the interaction with both 16 S rRNA and the mRNA target site.
J Mol Biol 1988 Nov 20
PMID:Translational regulation of the spc operon in Escherichia coli. Identification and structural analysis of the target site for S8 repressor protein. 246 92


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