Dimerization allows DNA target site recognition by the NarL response regulator

Maris, A.E.; Sawaya, M.R.; Kaczor-Grzeskowiak, Maria; Jarvis, Michael R.; Bearson, Shawn M.D.; Kopka, Mary L.; Schröder, Imke; Gunsalus, Robert P.; Dickerson, Richard E.

doi:10.1038/nsb845

Cited by 151 publications

(205 citation statements)

References 40 publications

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“…nt, nucleotide. of the two domains exposing critical DNA binding elements in the C-terminal domain (36). By analogy with the NarL activation model, we can speculate that unphosphorylated DesR dimerizes in solution by its C-terminal domain, whereas both N-terminal regulatory domains act as inhibitors of DNA binding by steric hindrance.…”

Section: Discussionmentioning

confidence: 99%

Bacillus subtilis DesR Functions as a Phosphorylation-activated Switch to Control Membrane Lipid Fluidity

Cybulski

Solar

Craig

et al. 2004

Journal of Biological Chemistry

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The Des pathway of Bacillus subtilis regulates the synthesis of the cold-shock induced membrane-bound enzyme ⌬5-fatty acid desaturase (⌬5-Des). A central component of the Des pathway is the response regulator, DesR, which is activated by a membrane-associated kinase, DesK, in response to a decrease in membrane lipid fluidity. Despite genetic and biochemical studies, specific details of the interaction between DesR and the DNA remain unknown. In this study we show that only the phosphorylated form of protein DesR is able to bind to a regulatory region immediately upstream of the promoter of the ⌬5-Des gene (Pdes). Phosphorylation of the regulatory domain of dimeric DesR promotes, in a cooperative fashion, the hierarchical occupation of two adjacent, non-identical, DesR-P DNA binding sites, so that there is a shift in the equilibrium toward the tetrameric active form of the response regulator. Subsequently, this phosphorylation signal propagation leads to the activation of the des gene through recruitment of RNA polymerase to Pdes. This is the first dissected example of a transcription factor functioning as a phosphorylationactivated switch for a cold-shock gene, allowing the cell to optimize the fluidity of membrane phospholipids.All organisms must communicate with their environment to survive. Bacterial cells monitor external conditions and process this information to give the most appropriate response. Twocomponent regulatory systems have emerged as a paradigm for adaptive responses. In its simplest form, a two-component system contains a sensor (histidine kinase) and a response regulator (often a transcriptional activator) (1). Changes in the environment result in phosphorylation of the sensor followed by transphosphorylation onto the response regulator (1, 2). Although kinase-response regulator pairs of this type were frequently reported as governors of a wide variety of pathways in response to a myriad of signals (3-5), the requirement of this system to control gene expression during cold-shock has only recently been discovered in Bacillus subtilis (6) and cyanobacteria (7).Cold shock is a stress condition that adversely affects the growth of poikilothermic organisms, such as bacteria and plants. Thus, understanding the mechanisms by which these organisms perceive low temperature signals and transmit this information to the cellular machinery to activate adaptive responses is of fundamental importance to biology.Bacteria and most (if not all) poikilothermic organisms have to remodel the membrane lipid composition to survive at low temperatures. B. subtilis (6) and Synechocystis (7) respond to a decrease in the ambient growth temperature by introducing double bonds into the acyl chains of their membrane phospholipids by membrane-bound acyl lipid desaturases. This postsynthetic modification of the saturated acyl chains seems to be designed to ameliorate the effect of temperature changes on the physical state of membrane phospholipids. In Synechocystis it was found that inactivation of two histidine kinases mod...

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Section: Discussionmentioning

confidence: 99%

Bacillus subtilis DesR Functions as a Phosphorylation-activated Switch to Control Membrane Lipid Fluidity

Cybulski

Solar

Craig

et al. 2004

Journal of Biological Chemistry

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“…35) as a structural template, we found that all of the top 20 DI pairs are true contacts (red bonds in Fig. 4A).…”

mentioning

confidence: 91%

Direct-coupling analysis of residue coevolution captures native contacts across many protein families

Morcos

Pagnani

Lunt

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

1,352

2,225

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The similarity in the three-dimensional structures of homologous proteins imposes strong constraints on their sequence variability. It has long been suggested that the resulting correlations among amino acid compositions at different sequence positions can be exploited to infer spatial contacts within the tertiary protein structure. Crucial to this inference is the ability to disentangle direct and indirect correlations, as accomplished by the recently introduced direct-coupling analysis (DCA). Here we develop a computationally efficient implementation of DCA, which allows us to evaluate the accuracy of contact prediction by DCA for a large number of protein domains, based purely on sequence information. DCA is shown to yield a large number of correctly predicted contacts, recapitulating the global structure of the contact map for the majority of the protein domains examined. Furthermore, our analysis captures clear signals beyond intradomain residue contacts, arising, e.g., from alternative protein conformations, ligand-mediated residue couplings, and interdomain interactions in protein oligomers. Our findings suggest that contacts predicted by DCA can be used as a reliable guide to facilitate computational predictions of alternative protein conformations, protein complex formation, and even the de novo prediction of protein domain structures, contingent on the existence of a large number of homologous sequences which are being rapidly made available due to advances in genome sequencing.statistical sequence analysis | residue-residue covariation | contact map prediction | maximum-entropy modeling

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“…The extract (yellow syrup) was purified twice by silica gel chromatography (silica gel 60N, 63-210 mm; Kanto Chemical, Tokyo, Japan; first: 1.5 10 cm gel with hexane/acetone (1:1), second 1.5 20 cm gel with hexane/EtOAc (1:4)) to obtain 282 mg of 25-O-methyl-FD-892. 1 H and 13 C NMR spectra were taken by DRX-500 (Bruker) or ECX500 (JEOL) spectrometer, and FAB-MS spectra were measured by JMS-700 (JEOL). The assignments of and SpeI, and the resulting DNA fragment was inserted into the corresponding site of pCYP-camAB [44] to obtain pGfsF-camAB.…”

Section: Methodsmentioning

confidence: 99%

“…A pOJ446-based cosmid library with the randomly Sau3AI digested genome DNA of S. graminofaciens A-8890 was constructed and further screened by hybridization with the DIG-labeled KS DNAs. As a result, one of the KS gene probes gave a gene cluster containing total 69 kbp of five modular type I PKS genes (gfsA, B, C, D, and E), a cytochrome P450 monooxygenase gene (gfsF), a methyltransferase gene (gfsG), and a NarL family two-component response regulator gene (gfsR), [13] which could be activated by a certain sensor protein in the microorganism ( Figure 1 and Table 1). There is no open-reading frame (ORF) over 1 kbp outside of the gene cluster; this suggests that this gene cluster is one operon for the biosynthesis of this particular polyketide.…”

Section: Identification Of the Gene Clustermentioning

confidence: 99%

Cloning and Characterization of the Biosynthetic Gene Cluster of 16‐Membered Macrolide Antibiotic FD‐891: Involvement of a Dual Functional Cytochrome P450 Monooxygenase Catalyzing Epoxidation and Hydroxylation

et al. 2010

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FD‐891 is a 16‐membered cytotoxic antibiotic macrolide that is especially active against human leukemia such as HL‐60 and Jurkat cells. We identified the FD‐891 biosynthetic (gfs) gene cluster from the producer Streptomyces graminofaciens A‐8890 by using typical modular type I polyketide synthase (PKS) genes as probes. The gfs gene cluster contained five typical modular type I PKS genes (gfsA, B, C, D, and E), a cytochrome P450 gene (gfsF), a methyltransferase gene (gfsG), and a regulator gene (gfsR). The gene organization of PKSs agreed well with the basic polyketide skeleton of FD‐891 including the oxidation states and α‐alkyl substituent determined by the substrate specificities of the acyltransferase (AT) domains. To clarify the involvement of the gfs genes in the FD‐891 biosynthesis, the P450 gfsF gene was inactivated; this resulted in the loss of FD‐891 production. Instead, the gfsF gene‐disrupted mutant accumulated a novel FD‐891 analogue 25‐O‐methyl‐FD‐892, which lacked the epoxide and the hydroxyl group of FD‐891. Furthermore, the recombinant GfsF enzyme coexpressed with putidaredoxin and putidaredoxin reductase converted 25‐O‐methyl‐FD‐892 into FD‐891. In the course of the GfsF reaction, 10‐deoxy‐FD‐891 was isolated as an enzymatic reaction intermediate, which was also converted into FD‐891 by GfsF. Therefore, it was clearly found that the cytochrome P450 GfsF catalyzes epoxidation and hydroxylation in a stepwise manner in the FD‐891 biosynthesis. These results clearly confirmed that the identified gfs genes are responsible for the biosynthesis of FD‐891 in S. graminofaciens.

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Dimerization allows DNA target site recognition by the NarL response regulator

Cited by 151 publications

References 40 publications

Bacillus subtilis DesR Functions as a Phosphorylation-activated Switch to Control Membrane Lipid Fluidity

Bacillus subtilis DesR Functions as a Phosphorylation-activated Switch to Control Membrane Lipid Fluidity

Direct-coupling analysis of residue coevolution captures native contacts across many protein families

Cloning and Characterization of the Biosynthetic Gene Cluster of 16‐Membered Macrolide Antibiotic FD‐891: Involvement of a Dual Functional Cytochrome P450 Monooxygenase Catalyzing Epoxidation and Hydroxylation

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