Surface Salt Bridges Modulate DNA Wrapping by the Type II DNA-Binding Protein TF1

Grove, Anne

doi:10.1021/bi034551o

Cited by 14 publications

(24 citation statements)

References 47 publications

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“…The wrapping of DNA about the surface of other HU homologs has been shown to involve disruption of surface salt bridges, and the wrapping path proposed to be defined by a specific pattern of surface salt bridges (39,40). Such coupled disruption of surface salt bridges is experimentally evidenced by a very modest salt-dependence of binding.…”

Section: Resultsmentioning

confidence: 99%

DNA protection by histone-like protein HU from the hyperthermophilic eubacterium Thermotoga maritima

Mukherjee

Sokunbi

Grove

2008

Nucleic Acids Research

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In mesophilic prokaryotes, the DNA-binding protein HU participates in nucleoid organization as well as in regulation of DNA-dependent processes. Little is known about nucleoid organization in thermophilic eubacteria. We show here that HU from the hyperthermophilic eubacterium Thermotoga maritima HU bends DNA and constrains negative DNA supercoils in the presence of topoisomerase I. However, while binding to a single site occludes ∼35 bp, association of T. maritima HU with DNA of sufficient length to accommodate multiple protomers results in an apparent shorter occluded site size. Such complexes consist of ordered arrays of protomers, as revealed by the periodicity of DNase I cleavage. Association of TmHU with plasmid DNA yields a complex that is remarkably resistant to DNase I-mediated degradation. TmHU is the only member of this protein family capable of occluding a 35 bp nonspecific site in duplex DNA; we propose that this property allows TmHU to form exceedingly stable associations in which DNA flanking the kinks is sandwiched between adjacent proteins. We suggest that T. maritima HU serves an architectural function when associating with a single 35 bp site, but generates a very stable and compact aggregate at higher protein concentrations that organizes and protects the genomic DNA.

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Section: Resultsmentioning

confidence: 99%

DNA protection by histone-like protein HU from the hyperthermophilic eubacterium Thermotoga maritima

Mukherjee

Sokunbi

Grove

2008

Nucleic Acids Research

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“…Despite the similar effects of N73A, K91A, and K94A on protein stability, K91A resulted in a much greater loss of DNA binding affinity than N73A or K94A ( Table 3), suggesting that K91 may also participate directly in DNA binding. In a growing number of DNA binding proteins, it has been observed that salt bridges formed near the DNA binding surface can become disrupted during DNA binding so that the cationic side chain can be free to interact with DNA (23,57,62). A similar process may occur with K91 in Fis.…”

Section: Discussionmentioning

confidence: 99%

Common and Variable Contributions of Fis Residues to High-Affinity Binding at Different DNA Sequences

et al. 2006

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Fis is a nucleoid-associated protein that interacts with poorly related DNA sequences with a high degree of specificity. A difference of more than 3 orders of magnitude in apparent K d values was observed between specific (K d , ϳ1 to 4 nM) and nonspecific (K d , ϳ4 M) DNA binding. To examine the contributions of Fis residues to the high-affinity binding at different DNA sequences, 13 alanine substitutions were generated in or near the Fis helix-turn-helix DNA binding motif, and the resulting proteins were purified. In vitro binding assays at three different Fis sites (fis P II, hin distal, and attR) revealed that R85, T87, R89, K90, and K91 played major roles in high-affinity DNA binding and that R85, T87, and K90 were consistently vital for binding to all three sites. Other residues made variable contributions to binding, depending on the binding site. N84 was required only for binding to the attR Fis site, and the role of R89 was dramatically altered by the attR DNA flanking sequence. The effects of Fis mutations on fis P II or hin distal site binding in vitro generally correlated with their abilities to mediate fis P repression or DNA inversion in vivo, demonstrating that the in vitro DNAbinding effects are relevant in vivo. The results suggest that while Fis is able to recognize a minimal common set of DNA sequence determinants at different binding sites, it is also equipped with a number of residues that contribute to the binding strength, some of which play variable roles.Fis (factor for inversion stimulation) is the most abundant nucleoid-associated protein during the logarithmic growth phase in rapidly growing Escherichia coli cells (3, 6). However, during mid-to late-logarithmic growth, the intracellular levels of Fis decrease over 500-fold and become nearly imperceptible during the stationary phase. The impact of Fis on cell physiology is widespread. As a nucleoid-associated protein, it is able to interact with a large number of DNA sites to alter DNA topology (66,67). Numerous genes are subject to positive or negative regulation by Fis directly or indirectly (6,10,19,22,30,49,56,61,(73)(74)(75). In the case of the promoters rrnB P1 and proP P2, Fis binding to sites centered at positions Ϫ71 and Ϫ41, respectively, directly stimulates transcription by contacting the C-terminal domain of the RNA polymerase ␣ subunit (␣-CTD) (9, 43). In the case of the fis promoter (fis P), Fis binding to sites I and II, centered at positions ϩ25 and Ϫ44, respectively, negatively autoregulates the fis operon by hindering RNA polymerase binding (6, 50, 58). As its name suggests, Fis also stimulates site-specific DNA inversion involving the Hin, Gin, and Cin family of recombinases (24,32,35). During Hin-mediated DNA inversion, Fis binds to two high-affinity DNA sites (hin-proximal and -distal sites) in a 60-bp recombinational enhancer region and interacts with two DNA-bound Hin recombinases to form a nucleoprotein complex intermediate required for efficient DNA strand cleavage and inversion (25,33). Bacteriophage DNA excis...

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“…HU proteins have been reported to interact with DNA with binding sites of between 9 and 42 bp (57). It has been suggested that variations in HU-binding site lengths are determined by the presence or absence of amino acids capable of forming salt bridges distal to sites of kinking (58). In HU homologs with shorter binding sites, K3 is proposed to form a salt bridge with D26.…”

Section: Discussionmentioning

confidence: 99%

DNA organization by the apicoplast-targeted bacterial histone-like protein of Plasmodium falciparum

Ram

Naik

Ganguli

et al. 2008

Nucleic Acids Research

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Apicomplexans, including the pathogens Plasmodium and Toxoplasma, carry a nonphotosynthetic plastid of secondary endosymbiotic origin called the apicoplast. The P. falciparum apicoplast contains a 35 kb, circular DNA genome with limited coding capacity that lacks genes encoding proteins for DNA organization and replication. We report identification of a nuclear-encoded bacterial histone-like protein (PfHU) involved in DNA compaction in the apicoplast. PfHU is associated with apicoplast DNA and is expressed throughout the parasite's intra-erythocytic cycle. The protein binds DNA in a sequence nonspecific manner with a minimum binding site length of ∼27 bp and a Kd of ∼63 nM and displays a preference for supercoiled DNA. PfHU is capable of condensing Escherichia coli nucleoids in vivo indicating its role in DNA compaction. The unique 42 aa C-terminal extension of PfHU influences its DNA condensation properties. In contrast to bacterial HUs that bend DNA, PfHU promotes concatenation of linear DNA and inhibits DNA circularization. Atomic Force Microscopic study of PfHU–DNA complexes shows protein concentration-dependent DNA stiffening, intermolecular bundling and formation of DNA bridges followed by assembly of condensed DNA networks. Our results provide the first functional characterization of an apicomplexan HU protein and provide additional evidence for red algal ancestry of the apicoplast.

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Surface Salt Bridges Modulate DNA Wrapping by the Type II DNA-Binding Protein TF1

Cited by 14 publications

References 47 publications

DNA protection by histone-like protein HU from the hyperthermophilic eubacterium Thermotoga maritima

DNA protection by histone-like protein HU from the hyperthermophilic eubacterium Thermotoga maritima

Common and Variable Contributions of Fis Residues to High-Affinity Binding at Different DNA Sequences

DNA organization by the apicoplast-targeted bacterial histone-like protein of Plasmodium falciparum

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