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PubMed2006

Kinetics of carboplatin-DNA binding in genomic DNA and bladder cancer cells as determined by accelerator mass spectrometry.

Cisplatin and carboplatin are platinum-based drugs that are widely used in cancer chemotherapy. The cytotoxicity of these drugs is mediated by platinum-DNA monoadducts and intra- and interstrand diadducts, which are formed following uptake of the drug into the nucleus of cells. The pharmacodynamics of carboplatin display fewer side effects than for cisplatin, albeit with less potency, which may be due to differences in rates of DNA adduct formation. We report the use of accelerator mass spectrometry (AMS), a sensitive detection method often used for radiocarbon quantitation, to measure both the kinetics of [14C]carboplatin-DNA adduct formation with genomic DNA and drug uptake and DNA binding in T24 human bladder cancer cells. Only carboplatin-DNA monoadducts contain radiocarbon in the platinated DNA, which allowed for calculation of kinetic rates and concentrations within the system. The percent of radiocarbon bound to salmon sperm DNA in the form of monoadducts was measured by AMS over 24 h. Knowledge of both the starting concentration of the parent carboplatin and the concentration of radiocarbon in the DNA at a variety of time points allowed calculation of the rates of Pt-DNA monoadduct formation and conversion to toxic cross-links. Importantly, the rate of carboplatin-DNA monoadduct formation was approximately 100-fold slower than that reported for the more potent cisplatin analogue, which may explain the lower toxicity of carboplatin. T24 human bladder cancer cells were incubated with a subpharmacological dose of [14C]carboplatin, and the rate of accumulation of radiocarbon in the cells and nuclear DNA was measured by AMS. The lowest concentration of radiocarbon measured was approximately 1 amol/10 microg of DNA. This sensitivity may allow the method to be used for clinical applications.

Antineoplastic Agents↗

PubMed2026

Activity of natural single nucleotide variants of human alkyladenine DNA glycosylase AAG R145H, G163S and R197C involved in DNA binding.

Alkyladenine DNA glycosylase (AAG) is a critical enzyme in the base excision repair (BER) pathway that safeguards genome integrity by removing structurally diverse alkylated and deaminated purine lesions from DNA. It serves as a primary defense against alkylation-induced mutations, which are linked to cancer development, chronic inflammation, and neurodegenerative diseases. Single nucleotide variants (SNVs) in the gene coding region have the potential to alter the enzyme's functionality, potentially modulating the repair capacity and affecting response and prognosis following chemoradiotherapy. In our study, we investigated three SNVs that lead to amino acid class changes in regions involved in DNA substrate coordination: R145H, G163S, and R197C using an in vitro approach. Using biochemical assays and molecular dynamics simulations, we evaluated the thermal stability, DNA binding affinity, and glycosylase activity of AAG variants toward hypoxanthine (Hx) and 1,N6-ethenoadenosine (εA) containing substrates. The G163S variant showed reduced thermal stability due to the conformational strain in the β-hairpin loop that intercalates in DNA, but retained εA excision activity comparable to that of wild-type AAG, while losing activity against Hx-containing DNA. The R197C variant had a four-fold reduction in DNA binding affinity for both substrates, and was catalytically inactive, unable to excise either damaged bases. This loss of function correlated with the rearrangement of the 201-210 loop and the reorientation of Arg-201 and Arg-207, which disrupts critical DNA contacts. However, the R145H variant retained near-wild-type thermal stability and activity on both substrates, despite bioinformatic predictions of deleterious effect. Molecular dynamics simulations revealed variant-specific structural disruptions. The data obtained underscore the importance of experimental validation in assessing the functional impact SNVs.

DNA Glycosylases↗

PubMed2000

The biochemical characterization of the DNA binding activity of pKi67.

Ki67 is only expressed in the nucleus of cycling cells. While it is employed as an operational marker of proliferation, little is known of the biochemical properties of this large protein. Using an immunoaffinity strategy for purification of pKi67, this study has shown that it can form higher-order complexes and can bind to DNA cellulose in vitro. No other co-purifying proteins could be identified, strongly suggesting that the DNA binding activity is an inherent property of pKi67. Using an electromobility shift assay, the affinity of pKi67 was shown using a range of different forms of DNA as competitors. Single-stranded DNA was the poorest competitor, followed by double-stranded DNA, with supercoiled DNA being the best competitor. In addition, it was found that purified pKi67 has a preference for AT-rich DNA. The DNA binding domain is mapped to the C-terminal domain of pKi67, and recombinant protein from the terminal 321 residues of pKi67 can bind DNA in vitro. GFP constructs from this domain were used to map regions that could target nucleolar localization and allow DNA binding. Finally, it was found that over-expression of the C-terminal 321 residues in cells induced chromatin disruption and apoptosis. These data provide strong evidence that pKi67 has a novel DNA binding activity within the C-terminal domain and that this protein can influence chromatin structure.

Apoptosis↗

PubMed1991

Redox manipulation of DNA binding activity and BuGR epitope reactivity of the glucocorticoid receptor.

Treatment of the transformed glucocorticoid receptor with hydrogen peroxide promotes the formation of disulfide bonds and inhibits the ability of the receptor to bind to DNA (Tienrungroj, W., Meshinchi, S., Sanchez, E. R., Pratt, S. E., Grippo, J. F., Holmgren, A., and Pratt, W. B. (1987) J. Biol. Chem. 262, 6992-7000). It has not been determined whether the inhibition of DNA binding activity is due to disulfide bonds formed within the DNA binding domain or between the DNA binding domain and another region of the receptor. In this paper, we examined the ability of hydrogen peroxide to inactivate the DNA binding activity of the mouse glucocorticoid receptor. We show that inhibition of DNA binding activity caused by hydrogen peroxide can be accounted for entirely by the formation of disulfide bonds between cysteine residues lying within the 15-kDa tryptic fragment containing the DNA binding domain of the receptor. Reversal of the peroxide-induced inactivation of DNA binding activity requires both zinc and a thiol-disulfide exchange reagent, such as dithiothreitol. Peroxide also eliminates recognition of the intact receptor and the 15-kDa tryptic fragment by the BuGR monoclonal antibody, and the reactivity of the BuGR epitope is restored by reduction without a requirement for zinc. Pretreatment of the receptor with methyl methanethiosulfonate inhibits much of the peroxide-mediated inactivation of the BuGR epitope but pretreatment with N-ethylmaleimide does not. Similarly, DNA binding activity of the receptor is inhibited by methyl methanethiosulfonate but not by N-ethylmaleimide. These results are consistent with the proposal that peroxide promotes the formation of disulfide bonds between thiols that lie spatially close to one another in the 15-kDa tryptic fragment, resulting in rapid elimination of zinc. Restoration of the zinc finger structure restores DNA-binding activity but restoration of the BuGR epitope requires only reduction without restoration of the zinc fingers.

Animals↗

PubMed1979

Gene D5 product of bacteriophage T5: DNA-binding protein affecting DNA replication and late gene expression.

Gene D5 is not only necessary for replication of bacteriophage T5 DNA and for shutoff of expression of some early genes, but has been found to be necessary also for the expression of late T5 genes. The polypeptide product of gene D5 has been identified, an intragenic map of gene D5 has been constructed, and the direction of transcription of gene D5 has been established. The polypeptide coded by gene D5 has been shown to be a DNA-binding protein with affinity for both double- and single-stranded DNA.

Coliphages↗

PubMed2005

Doxazosin induces apoptosis in LNCaP prostate cancer cell line through DNA binding and DNA-dependent protein kinase down-regulation.

Doxazosin is a quinazoline-based compound acting as an alpha-1-adrenergic inhibitor shown to induce apoptosis in prostate cancer cell lines via an alpha-1-adrenergic receptor-independent mechanism. To better understand the mechanism of doxazosin-induced apoptosis in prostate cancer, we performed cDNA microarray to analyze gene expression changes produced by doxazosin in the androgen-dependent human prostate cancer cell line, LNCaP. We found that 70 and 92 genes were deregulated after 8 and 24 h of doxazosin treatment, respectively. These genes are involved in several cellular processes such as cell-cycle regulation, cell adhesion and signal transduction pathways. Strikingly, we found that doxazosin induces deregulation of genes implicated in DNA replication and repair, such as GADD45A, XRCC5 and PRKDC. These facts, together with the demonstration of the ability of doxazosin to bind DNA, allowed us to propose a novel mechanism of action for doxazosin in prostate cancer cells that implies DNA-damage mediated apoptosis by down-regulation of XRCC5 and PRKDC genes.

Adrenergic alpha-Antagonists↗

PubMed2004

DNA-mediated assembly of weakly interacting DNA-binding protein subunits: in vitro recruitment of phage 434 repressor and yeast GCN4 DNA-binding domains.

The specificity of DNA-mediated protein assembly was studied in two in vitro systems, based on (i) the DNA-binding domain of bacteriophage 434 repressor cI (amino acid residues 1-69), or (ii) the DNA-binding domain of the yeast transcription factor GCN4, (amino acids 1-34) and their respective oligonucleotide cognates. In vivo, both of these peptides are part of larger protein molecules that also contain dimerization domains, and the resulting dimers recognize cognate palindromic DNA sequences that contain two half-sites of 4 bp each. The dimerization domains were not included in the peptides tested, so in solution-in the presence or absence of non-cognate DNA oligonucleotides-these molecules did not show appreciable dimerization, as determined by pyrene excimer fluorescence spectroscopy and oxidative cross-linking monitored by mass spectrometry. Oligonucleotides with only one 4 bp cognate half-site were able to initiate measurable dimerization, and two half-sites were able to select specific dimers even from a heterogeneous pool of molecules of closely related specificity (such as DNA-binding domains of the 434 repressor and their engineered mutants that mimic the binding helix of the related P22 phage repressor). The fluorescent technique allowed us to separately monitor the unspecific, ionic interaction of the peptides with DNA which produced a roughly similar signal in the case of both cognate and non-cognate oligonucleotides. But in the former case, a concomitant excimer fluorescence signal showed the formation of correctly positioned dimers. The results suggest that DNA acts as a highly specific template for the recruitment of weakly interacting protein molecules that can thus build up highly specific complexes.

Amino Acid Sequence↗

PubMed2006

Characterization of DNA-binding specificity and analysis of binding sites of the Pseudomonas aeruginosa global regulator, Vfr, a homologue of the Escherichia coli cAMP receptor protein.

Vfr, a global regulator of Pseudomonas aeruginosa virulence factors, is a homologue of the Escherichia coli cAMP receptor protein, CRP. Vfr is 91% similar to CRP and maintains many residues important for CRP to bind cAMP, bind DNA, and interact with RNA polymerase at target promoters. While vfr can complement an E. coli crp mutant in beta-galactosidase production, tryptophanase production and catabolite repression, crp can only complement a subset of Vfr-dependent phenotypes in P. aeruginosa. Using specific CRP binding site mutations, it is shown that Vfr requires the same nucleotides as CRP for optimal transcriptional activity from the E. coli lac promoter. In contrast, CRP did not bind Vfr target sequences in the promoters of the toxA and regA genes. Footprinting analysis revealed Vfr protected sequences upstream of toxA, regA, and the quorum sensing regulator lasR, that are similar to but significantly divergent from the CRP consensus binding sequence, and Vfr causes similar DNA bending to CRP in bound target sequences. Using a preliminary Vfr consensus binding sequence deduced from the Vfr-protected sites, Vfr target sequences were identified upstream of the virulence-associated genes plcN, plcHR, pbpG, prpL and algD, and in the vfr/orfX, argH/fimS, pilM/ponA intergenic regions. From these sequences the Vfr consensus binding sequence, 5'-ANWWTGNGAWNY : AGWTCACAT-3', was formulated. This study suggests that Vfr shares many of the same functions as CRP, but has specialized functions, at least in terms of DNA target sequence binding, required for regulation of a subset of genes in its regulon.

ADP Ribose Transferases↗

PubMed2003

The ATP-dependent Lon protease of Mus musculus is a DNA-binding protein that is functionally conserved between yeast and mammals.

The ATP-dependent Lon protease is a multi-functional enzyme that is conserved from archae to mammalian mitochondria, which not only degrades protein substrates but also binds DNA. As a starting point toward understanding Lon function in development, the mouse Lon cDNA was cloned and the encoded protein was characterized in cultured mammalian cells, in yeast and in vitro. Mouse Lon shows 87, 40 and 33% amino acid similarity with the human, yeast and bacterial homologs, respectively. Expression of a single mouse Lon transcript is detected in liver>heart>kidney>testis and is present during early embryonic development. Endogenous as well as transiently overexpressed mouse Lon co-localize with mitochondrial markers and have half-lives greater than 24 h as determined by pulse-chase studies. Enzymatically active mouse Lon that hydrolyses ATP and degrades protein and peptide substrates in an ATP-dependent manner also specifically binds to single-stranded but not to double-stranded DNA oligonucleotides. We propose that binding to TG-rich DNA sequences has been conserved between the mouse and human proteins. In addition, the evolutionary conservation of mitochondrial Lon function is demonstrated by the ability of mouse Lon to substitute for the yeast protein in vivo.

ATP-Dependent Proteases↗

PubMed1996

The DNA-binding protein Hdf1p (a putative Ku homologue) is required for maintaining normal telomere length in Saccharomyces cerevisiae.

In mammalian cells, the Ku autoantigen is an end- binding DNA protein required for the repair of DNA breaks [Troelstra, C. and Jaspers, N.G.J. (1994) Curr. Biol., 4, 1149- 1151]. A yeast gene (HDF1) encoding a putative homologue of the 70 kDa subunit of Ku has recently been identified [Feldmann, H. and Winnacker, E. L. (1993) J. Biol. Chem., 268, 12895- 12900]. We find that hdf1 mutant strains have substantially shorter telomeres than wild-type strains. We speculate that Hdf1p may bind the natural ends of the chromosome, in addition to binding to the ends of broken DNA molecules. Strains with both an hdf1 mutation and a mutation in TEL 1 (a gene related to the human ataxia telangiectasia gene) have extremely short telomeres and grow slowly.

Antigens, Nuclear↗

PubMed1994

Stimulation of site-specific topoisomerase II-mediated DNA cleavage by an N-methylpyrrolecarboxamide-anilinoacridine conjugate: relation to DNA binding.

The DNA binding properties and effects on topoisomerase II of MePyGA, an anilinoacridine derivative bearing an N-methylpyrrolecarboxamide unit at position 1', have been compared with those of its precursor glycylanilinoacridine and the structurally related antileukaemic drug amsacrine. Electric linear dichroism spectroscopy reveals that MePyGA intercalates its acridine chromophore between DNA base pairs with a preference for GC-rich sequences, whereas both its structural analogue lacking the N-methylpyrrole unit and amsacrine intercalate into DNA without any strong sequence preference. The effects of the test drug on the catalytic activities of topoisomerase II were studied in vitro using purified calf thymus enzyme and 32P-labeled DNA. MePyGA stabilizes the topoisomerase II-DNA covalent complex and stimulates the cutting of DNA at a subset of preexisting topoisomerase II cleavage sites. The removal of the N-methylpyrrole unit abolishes both the GC-preferential binding to DNA and the topoisomerase II-mediated DNA cleavage. MePyGA and amsacrine stimulate the cleavage of DNA by topoisomerase II at different places: cleavage stimulated by amsacrine is consistent with the expected adenine requirement at position +1 whereas the predominant sites of DNA cleavage stimulated by MePyGA contain a cytosine at position +/- 1. This is the first instance where an anilinoacridine derivative differing only by the nature of the substituent at position 1' has been found to affect the catalytic activity of topoisomerase II differently. The spectroscopic and biochemical data lead to the conclusion that two functional domains can be identified in MePyGA: its anilino group can be regarded as a skeletal core to which are connected (i) the tricyclic acridine moiety which represents the DNA-binding domain and (ii) the N-methylpyrrole moiety which constitutes the topoisomerase II-targeted domain. The structure of the substituent at position 1' of the anilinoacridine chromophore evidently determines the location of the sites of DNA cleavage by topoisomerase II. These findings provide guidance for the synthesis and development of new topoisomerase II-targeted antitumor anilinoacridine derivatives.

Acridines↗

PubMed2002

The ability of cystamine to bind DNA.

The DNA-binding properties of cystamine compared with natural occurring polyamines have been studied in vitro by means of ethidium bromide displacement assays, studies of DNA thermal stability and analyses of DNA-B/DNA-A transition. While the first two methods did not put in evidence any peculiar property in the binding capability of cystamine, CD studies showed the interesting ability of cystamine to shift the equilibrium B/A-DNA towards the B-form. In the same experimental conditions spermine and spermidine induced the A form of DNA, instead putrescine and cadaverine did not show any particular activity. The ability of cystamine to bind DNA, as shown also by its DNA radioprotective capability, might be important in chromatin condensation and stabilization, and might be a cause of the antiviral activity observed by some authors.

Animals↗

PubMed2005

Synthesis, characterization and DNA-binding properties of rac-[Ru(5,6-dmp)2(dppz)]2+--enantiopreferential DNA binding and co-ligand promoted exciton coupling.

The new mixed ligand complex [Ru(5,6-dmp)2(dppz)]Cl2 [5,6-dmp = 5,6-dimethyl-1,10-phenanthroline, dppz = dipyrido[3,2-a:2',3'-c]phenazine] has been isolated and its DNA-binding properties studied by employing UV-visible (UV-Vis), steady-state and time-resolved emission and circular dichroism spectral methods, viscometry, thermal denaturation and cyclic/differential pulse voltammetric techniques. The complex acts as a 'molecular light-switch' on binding to DNA, but the enhancement in emission intensity is only 75% of that of the parent complex [Ru(phen)2(dppz)]2+ (phen = 1,10-phenanthroline). The emission decay curves and quenching studies suggest two different DNA-binding modes both involving intercalation of the dppz ligand of [Ru(5,6-dmp)2(dppz)]Cl2. The characteristic red-shift of the induced CD signal, which is not observed for the phen analogue, arises from exciton coupling. The hydrophobicity and polarizability of 5,6-dmp co-ligand strongly favour the formation of a stable structural and electronic scaffold on the DNA surface for the unbound molecules to couple with the DNA-bound complexes facilitating spontaneous assembly of novel extended molecular aggregates using DNA as a helical nanotemplate. This observation is consistent with the shift in Ru(II)/Ru(III) redox potential to more positive values with a dramatic drop in peak current on binding of the 5,6-dmp complex to calf thymus (CT) DNA. Equilibrium dialysis experiments monitored by CD spectroscopy unambiguously reveal the preferential binding of the delta-enantiomer to the right-handed calf thymus (CT) DNA. The 5,6-dmp complex exhibits preferential binding to [d(AT)6]2 over [d(GC)6]2 and the complex aggregates formed consist of six [Ru(5,6-dmp)2(dppz)]2+ cations per base pair of [d(AT)6]2; however, only one [Ru(phen)2(dppz)]2+ cation per base pair is involved in DNA binding.

Circular Dichroism↗

PubMed2002

Characterisation of the DNA-binding profile of barley HvCBF1 using an enzymatic method for rapid, quantitative and high-throughput analysis of the DNA-binding activity.

A rapid and quantitative DNA-binding assay was developed based on the translational fusion of a DNA-binding protein (DBP) with a Neocallimastix patriciarum beta-1,4-D-glucanase, CelD. CelD releases a fluorescent 4-methylumbelliferyl product from 4-methylumbelliferyl cellobioside. This hydrolysis activity was used to quantify the amount of DBP-CelD bound to an immobilised biotin-labelled target sequence. The DNA-binding assay can be performed in a 96-well plate format for high- throughput analysis of putative DBPs. This method was applied to analysis of the binding properties and sequence selectivity of a cold-inducible transcription factor HvCBF1 from barley containing an AP2 DNA-binding domain. A base-scanning approach using degenerate oligonucleotide probes was employed for rapid identification of the conserved core motif of the HvCBF1 binding site. Quantitative analysis of the binding site of HvCBF1 using systematic base substitution revealed that a (G/a) (C/t)CGAC sequence was sufficient to constitute a functional motif, where the lower-case letters represent less efficient bases. The method enables us to provide accurate and quantitative data on a comprehensive DNA-binding profile for a cold-inducible AP2 transcription factor as well as information on environmental parameters potentially influencing the DNA-binding activity. The accurate binding sequence data facilitate identification of candidate genes regulated by HvCBF1 from genome sequence databases.

Base Sequence↗

PubMed1998

Evidence for two modes of cooperative DNA binding in vivo that do not involve direct protein-protein interactions.

BACKGROUND: The promoter regions of most eukaryotic genes contain binding sites for more than one transcriptional activator and these activators often bind cooperatively to promoters. The most common type of cooperativity is supported by direct protein-protein interactions. Recent studies have shown that proteins that do not specifically interact with one another can bind cooperatively to chromatin in vitro. probably by the localized destabilization of nucleosome structure by one factor, facilitating binding of another to a nearby site. This mechanism does not require that the transcription factors have activation domains. We have examined whether this phenomenon occurs in vivo. RESULTS: Unrelated non-interacting proteins can bind DNA cooperatively in yeast cells; this cooperative binding can contribute significantly to transcriptional activation, does not require that both factors have activation domains and is only operative over relatively short distances. In addition to this 'short-range' mechanism, unrelated non-interacting proteins can bind cooperatively to sites separated by hundreds of base pairs, so long as both have potent activation domains. CONCLUSION: Cooperative binding of transcription factors in vivo can occur by several mechanisms, some of which do not require direct protein-protein interactions and which cannot be detected in vitro using naked DNA templates. These findings must be taken into account when evaluating mechanisms for synergistic transcriptional activation.

Bacterial Proteins↗

PubMed1994

Determinants of affinity and mode of DNA binding at the carboxy terminus of the bacteriophage SPO1-encoded type II DNA-binding protein, TF1.

The role of the carboxy-terminal amino acids of the bacteriophage SPO1-encoded type II DNA-binding protein, TF1, in DNA binding was analyzed. Chain-terminating mutations truncating the normally 99-amino-acid TF1 at amino acids 96, 97, and 98 were constructed, as were missense mutations substituting cysteine, arginine, and serine for phenylalanine at amino acid 97 and tryptophan for lysine at amino acid 99. The binding of the resulting proteins to a synthetic 44-bp binding site in 5-(hydroxymethyl)uracil DNA, to binding sites in larger SPO1 [5-(hydroxymethyl)uracil-containing] DNA fragments, and to thymine-containing homologous DNA was analyzed by gel retardation and also by DNase I and hydroxy radical footprinting. We conclude that the C tail up to and including phenylalanine at amino acid 97 is essential for DNA binding and that the two C-terminal amino acids, 98 and 99, are involved in protein-protein interactions between TF1 dimers bound to DNA.

Amino Acid Sequence↗

PubMed2002

Iron and hydrogen peroxide detoxification properties of DNA-binding protein from starved cells. A ferritin-like DNA-binding protein of Escherichia coli.

The DNA-binding proteins from starved cells (Dps) are a family of proteins induced in microorganisms by oxidative or nutritional stress. Escherichia coli Dps, a structural analog of the 12-subunit Listeria innocua ferritin, binds and protects DNA against oxidative damage mediated by H(2)O(2). Dps is shown to be a Fe-binding and storage protein where Fe(II) oxidation is most effectively accomplished by H(2)O(2) rather than by O(2) as in ferritins. Two Fe(2+) ions bind at each of the 12 putative dinuclear ferroxidase sites (P(Z)) in the protein according to the equation, 2Fe(2+) + P(Z) --> [(Fe(II)(2)-P](FS)(Z+2) + 2H(+). The ferroxidase site (FS) bound iron is then oxidized according to the equation, [(Fe(II)(2)-P](FS)(Z+2) + H(2)O(2) + H(2)O --> [Fe(III)(2)O(2)(OH)-P](FS)(Z-1) + 3H(+), where two Fe(II) are oxidized per H(2)O(2) reduced, thus avoiding hydroxyl radical production through Fenton chemistry. Dps acquires a ferric core of approximately 500 Fe(III) according to the mineralization equation, 2Fe(2+) + H(2)O(2) + 2H(2)O --> 2Fe(III)OOH((core)) + 4H(+), again with a 2 Fe(II)/H(2)O(2) stoichiometry. The protein forms a similar ferric core with O(2) as the oxidant, albeit at a slower rate. In the absence of H(2)O(2) and O(2), Dps forms a ferrous core of approximately 400 Fe(II) by the reaction Fe(2+) + H(2)O + Cl(-) --> Fe(II)OHCl((core)) + H(+). The ferrous core also undergoes oxidation with a stoichiometry of 2 Fe(II)/H(2)O(2). Spin trapping experiments demonstrate that Dps greatly attenuates hydroxyl radical production during Fe(II) oxidation by H(2)O(2). These results and in vitro DNA damage assays indicate that the protective effect of Dps on DNA most likely is exerted through a dual action, the physical association with DNA and the ability to nullify the toxic combination of Fe(II) and H(2)O(2). In the latter process a hydrous ferric oxide mineral core is produced within the protein, thus avoiding oxidative damage mediated by Fenton chemistry.

Bacterial Proteins↗

PubMed2000

Bacillus subtilis LrpC is a sequence-independent DNA-binding and DNA-bending protein which bridges DNA.

Genetic evidence suggests that the Bacillus subtilis lrpC gene product participates in cell growth and sporulation. The purified LrpC protein, which has a predicted molecular mass of 16.4 kDa, is a tetramer in solution. LrpC binds with higher affinity ( K (app) approximately 80 nM) to intrinsically curved DNA than to non-curved DNA ( K (app) approximately 700 nM). DNase I footprinting and the supercoiling of relaxed circular plasmid DNA in the presence of topoisomerase I revealed that LrpC induces DNA bending and constrains DNA supercoils in vitro. The LrpC protein cooperatively increases DNA binding of the bona fide DNA-binding and DNA-bending protein Hbsu. LrpC forms inter- and intramolecular bridges on linear and supercoiled DNA molecules, resulting in a large network and DNA compactation. Collectively, these findings suggest that LrpC is an architectural protein and that its activities could provide a means to modulate DNA transactions.

Bacillus subtilis↗