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Repressor Protein (repressor + protein)
Selected AbstractsBinding properties of peptidic affinity ligands for plasmid DNA capture and detectionAICHE JOURNAL, Issue 2 2009Ying Han Abstract Peptides constructed from ,-helical subunits of the Lac repressor protein (LacI) were designed then tailored to achieve particular binding kinetics and dissociation constants for plasmid DNA purification and detection. Surface plasmon resonance was employed for quantification and characterization of the binding of double stranded Escherichia coli plasmid DNA (pUC19) via the lac operon (lacO) to "biomimics" of the DNA binding domain of LacI. Equilibrium dissociation constants (KD), association (ka), and dissociation rates (kd) for the interaction between a suite of peptide sequences and pUC19 were determined. KD values measured for the binding of pUC19 to the 47mer, 27mer, 16mer, and 14mer peptides were 8.8 ± 1.3 × 10,10 M, 7.2 ± 0.6 × 10,10 M, 4.5 ± 0.5 × 10,8 M, and 6.2 ± 0.9 × 10,6 M, respectively. These findings show that affinity peptides, composed of subunits from a naturally occurring operon,repressor interaction, can be designed to achieve binding characteristics suitable for affinity chromatography and biosensor devices. © 2008 American Institute of Chemical Engineers AIChE J, 2009 [source] Regulation of AmtR-controlled gene expression in Corynebacterium glutamicum: mechanism and characterization of the AmtR regulonMOLECULAR MICROBIOLOGY, Issue 2 2005Gabriele Beckers Summary AmtR, the master regulator of nitrogen control in Corynebacterium glutamicum, represses transcription of a number of genes during nitrogen surplus. Repression is released by an interaction of AmtR with signal transduction protein GlnK. As shown by pull-down assays and gel retardation experiments, only adenylylated GlnK, which is present in the cells during nitrogen limitation, is able to bind to AmtR. The AmtR regulon was characterized in this study by a combination of bioinformatics, transcriptome and proteome analyses. At least 33 genes are directly controlled by the repressor protein including those encoding transporters and enzymes for ammonium assimilation (amtA, amtB, glnA, gltBD), urea and creatinine metabolism (urtABCDE, ureABCEFGD, crnT, codA), a number of biochemically uncharacterized enzymes and transport systems (NCgl1099, NCgl1100, NCgl 1915,1918) as well as signal transduction proteins (glnD, glnK). For the AmtR regulon, an AmtR box has been defined which comprises the sequence tttCTATN6AtAGat/aA. Furthermore, the transcriptional organization of AmtR-regulated genes and operons was characterized. [source] Riboregulation by DsrA RNA: trans -actions for global economyMOLECULAR MICROBIOLOGY, Issue 4 2000MicroReview DsrA is an 87 nucleotide Escherichia coli RNA with extraordinary regulatory properties. The profound impact of its actions stems from DsrA regulating translation of two global transcription regulators, H-NS and RpoS (,s), by sequence-specific RNA,RNA interactions. H-NS is a major nucleoid-structuring and global repressor protein, and RpoS is the stationary phase and stress response sigma factor of RNA polymerase. DsrA changes its conformation to bind to these two different mRNA targets and thereby inhibits H-NS translation, while stimulating that of RpoS in a mechanistically distinct fashion. DsrA apparently binds both the start and the stop codons of hns mRNA and sharply decreases the mRNA half-life. DsrA also binds sequences in the 5,-untranslated leader region of rpoS mRNA, enhancing rpoS mRNA stability and RpoS translation. A cohort of genes, governed by H-NS repression and RpoS activation, are thus regulated. Low temperatures increase the levels of DsrA, with differential effects on H-NS and RpoS. Additionally, the RNA chaperone protein Hfq is involved with DsrA regulation, as well as with other small RNAs that also act on RpoS to co-ordinate stress responses. We address the possible functions of this genetic regulatory mechanism, as well as the advantages of using small RNAs as global regulators to orchestrate gene expression. [source] Allosteric transition pathways in the lactose repressor protein core domains: Asymmetric motions in a homodimerPROTEIN SCIENCE, Issue 11 2003Terence C. Flynn Abstract The crystal structures of lactose repressor protein (LacI) provide static endpoint views of the allosteric transition between DNA- and IPTG-bound states. To obtain an atom-by-atom description of the pathway between these two conformations, motions were simulated with targeted molecular dynamics (TMD). Strikingly, this homodimer exhibited asymmetric dynamics. All asymmetries observed in this simulation are reproducible and can begin on either of the two monomers. Asymmetry in the simulation originates around D149 and was traced back to the pre-TMD equilibrations of both conformations. In particular, hydrogen bonds between D149 and S193 adopt a variety of configurations during repetitions of this process. Changes in this region propagate through the structure via noncovalent interactions of three interconnected pathways. The changes of pathway 1 occur first on one monomer. Alterations move from the inducer-binding pocket, through the N-subdomain ,-sheet, to a hydrophobic cluster at the top of this region and then to the same cluster on the second monomer. These motions result in changes at (1) side chains that form an interface with the DNA-binding domains and (2) K84 and K84', which participate in the monomer,monomer interface. Pathway 2 reflects consequent reorganization across this subunit interface, most notably formation of a H74-H74rsquo; ,-stacking intermediate. Pathway 3 extends from the rear of the inducer-binding pocket, across a hydrogen-bond network at the bottom of the pocket, and transverses the monomer,monomer interface via changes in H74 and H74rsquo;. In general, intermediates detected in this study are not apparent in the crystal structures. Observations from the simulations are in good agreement with biochemical data and provide a spatial and sequential framework for interpreting existing genetic data. [source] NtSET1, a member of a newly identified subgroup of plant SET-domain-containing proteins, is chromatin-associated and its ectopic overexpression inhibits tobacco plant growthTHE PLANT JOURNAL, Issue 4 2001Wen-Hui Shen Summary The SET- and chromo-domains are recognized as signature motifs for proteins that contribute to epigenetic control of gene expression through effects on the regional organization of chromatin structure. This paper reports the identification of a novel subgroup of SET-domain-containing proteins in tobacco and Arabidopsis, which show highest homologies with the Drosophila position-effect-variegation repressor protein SU(VAR)3,9 and the yeast centromer silencing protein CLR4. The tobacco SET-domain-containing protein (NtSET1) was fused to the green fluorescence protein (GFP) that serves as a visual marker for localization of the recombinant protein in living cells. Whereas control GFP protein alone was uniformly dispersed within the nucleus and cytoplasm, the NtSET1-GFP fusion protein showed a non-uniform localization to multiple nuclear regions in interphase tobacco TBY2 cells. During mitosis, the NtSET1-GFP associated with condensed chromosomes with a non-random distribution. The NtSET1 thus appears to have distinct target regions in the plant chromatin. Overexpression of the NtSET1-GFP in transgenic tobacco inhibited plant growth, implicating the possible involvement of the NtSET1 in transcriptional repression of growth control genes through the formation of higher-order chromatin domains. [source] Structure of the apo form of the catabolite control protein A (CcpA) from Bacillus megaterium with a DNA-binding domainACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2007Rajesh Kumar Singh Crystal structure determination of catabolite control protein A (CcpA) at 2.6,Å resolution reveals for the first time the structure of a full-length apo-form LacI-GalR family repressor protein. In the crystal structures of these transcription regulators, the three-helix bundle of the DNA-binding domain has only been observed in cognate DNA complexes; it has not been observed in other crystal structures owing to its mobility. In the crystal packing of apo-CcpA, the protein,protein contacts between the N-terminal three-helix bundle and the core domain consisted of interactions between the homodimers that were similar to those between the corepressor protein HPr and the CcpA N-subdomain in the ternary DNA complex. In contrast to the DNA complex, the apo-CcpA structure reveals large subdomain movements in the core, resulting in a complete loss of contacts between the N-subdomains of the homodimer. [source] Physicochemical properties and distinct DNA binding capacity of the repressor of temperate Staphylococcus aureus phage ,11FEBS JOURNAL, Issue 7 2009Tridib Ganguly The repressor protein and cognate operator DNA of any temperate Staphylococcus aureus phage have not been investigated in depth, despite having the potential to enrich the molecular biology of the staphylococcal system. In the present study, using the extremely pure repressor of temperate Staphylococcus aureus phage ,11 (CI), we demonstrate that CI is composed of ,-helix and ,-sheet to a substantial extent at room temperature, possesses two domains, unfolds at temperatures above 39 °C and binds to two sites in the ,11 cI - cro intergenic region with variable affinity. The above CI binding sites harbor two homologous 15 bp inverted repeats (O1 and O2), which are spaced 18 bp apart. Several guanine bases located in and around O1 and O2 demonstrate interaction with CI, indicating that these 15 bp sites are used as operators for repressor binding. CI interacted with O1 and O2 in a cooperative manner and was found to bind to operator DNA as a homodimer. Interestingly, CI did not show appreciable binding to another homologous 15 bp site (O3) that was located in the same primary immunity region as O1 and O2. Taken together, these results suggest that ,11 CI and the ,11 CI,operator complex resemble significantly those of the lambdoid phages at the structural level. The mode of action of ,11 CI, however, may be distinct from that of the repressor proteins of , and related phages. [source] | |||||||||||||