DNA supercoiling promotes formation of a bent repression loop in lac DNA

Borowiec, James A.; Li, Zhang; Sasse-Dwight, Selina; Gralla, Jay D.

doi:10.1016/0022-2836(87)90513-4

Cited by 239 publications

(179 citation statements)

References 32 publications

Supporting

Mentioning

176

Contrasting

Order By: Relevance

“…Our analysis of the distribution of roll and twist angles in optimized loop structures shows that published permanganate footprinting profiles (22) are compatible with the antiparallel A1 configuration. Preliminary calculations for 452-bp negatively supercoiled DNA plasmids with a bound LacR tetramer further indicate that the optimal spacing for stable A1 loop formation is 162 bp when ⌬Lk ϭ Ϫ1, a result in accord with the gel observations of Krämer et al (30).…”

Section: Discussionmentioning

confidence: 61%

“…Addition of potassium permanganate, a probe for locally distorted DNA (21), to the wild-type LacRmediated O3-O1 loop reveals a hypersensitive base-pair region, H ϭ TTTAT, located Ϸ37 bp upstream of the O1 site (22). For DNA, TA steps are known to be sites of permanganate attack (23), and the configuration of such steps in the H region of the O3-O1 loop is apt to resemble the KMnO 4 -sensitive TA step (with high twist and negative roll) found in the crystal complex of DNA with the Epstein-Barr virus origin-binding protein (24).…”

Section: Resultsmentioning

confidence: 99%

“…For four of these, A1, A2, P1, and P2, the LacR is in its V-shaped conformation; for P1 E the LacR has its extended form. Employing a base-pair-level theory of sequence-dependent DNA elasticity, we have calculated the deformational free energy of the DNA in the complex for each type of loop and used the results to relate protein and DNA fine structure to a wide range of heretofore-unconnected experimental observations, including (i) loop stabilities (20,30), (ii) permanganate sensitivity (22), and (iii) DNase I cutting patterns (20). This work provides an understanding of the effects of chain length, base-pair sequence, protein binding, and supercoiling on DNA looping preferences and, in principle, allows one to design new ways to test specific looped structures in the laboratory.…”

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Modeling the Lac repressor-operator assembly: The influence of DNA looping on Lac repressor conformation

Swigon

Coleman

Olson³

2006

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

118

192

View full text Add to dashboard Cite

Repression of transcription of the Escherichia coli Lac operon by the Lac repressor (LacR) is accompanied by the simultaneous binding of LacR to two operators and the formation of a DNA loop. A recently developed theory of sequence-dependent DNA elasticity enables one to relate the fine structure of the LacR-DNA complex to a wide range of heretofore-unconnected experimental observations. Here, that theory is used to calculate the configuration and free energy of the DNA loop as a function of its length and base-pair sequence, its linking number, and the end conditions imposed by the LacR tetramer. The tetramer can assume two types of conformations. Whereas a rigid V-shaped structure is observed in the crystal, EM images show extended forms in which two dimer subunits are flexibly joined. Upon comparing our computed loop configurations with published experimental observations of permanganate sensitivities, DNase I cutting patterns, and loop stabilities, we conclude that linear DNA segments of short-to-medium chain length (50 -180 bp) give rise to loops with the extended form of LacR and that loops formed within negatively supercoiled plasmids induce the V-shaped structure.lac operon ͉ sequence-dependent DNA elasticity ͉ DNase I footprinting M any genetic processes are controlled by proteins that bind at separate, often widely spaced, sites on DNA and hold the intervening double helix in a loop (1-3). The classical example is the lac operon of Escherichia coli (4). The Lac repressor (LacR) is a tetrameric protein assembly that represses the expression of the lac operon by simultaneously binding to two DNA sites, i.e., operators, in the vicinity of the nucleotides at which transcription starts. The structure and elastic properties of DNA determine which spacings of the operators are optimal for functionality. For the lac operon, a change in spacing by five to six nucleotides can induce a 50-fold alteration in the efficiency of repression (5, 6).Although there is a large amount of literature on genetic and biochemical aspects of expression in the lac system, less is known about the actual configuration of the LacR-DNA loop assembly. In the crystalline state the two dimer subunits of LacR are joined to form a V (7, 8), and contact with DNA is made at the tips of each arm of the V (Fig. 1). On the other hand, electron microscopy and solution studies (9-12) indicate that the angle between the dimer subunits, i.e., the angle of aperture ␣, can vary. A change in ␣ affects the configuration of the DNA loop through its influence on the distance and orientation of the operators. We are here concerned with loops formed between the primary operator site O1 and the weaker auxiliary site O3. As each operator binds to the protein in one of two possible orientations, there are four distinct loop types that are analogous to those considered by Geanacopoulos et al. (13) in their treatment of DNA loops in the E. coli gal operon. We write A1, A2, P1, and P2 for these loop types, where the A and P refer to antiparallel and parallel ...

show abstract

Section: Discussionmentioning

confidence: 61%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Modeling the Lac repressor-operator assembly: The influence of DNA looping on Lac repressor conformation

Swigon

Coleman

Olson³

2006

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

118

192

View full text Add to dashboard Cite

show abstract

“…Specific f lexibly kinked DNA structures have been observed in all-atom molecular dynamics simulations of a 94-bp DNA minicircle (F. Lankas, R. Lavery, and J. H. Maddocks, personal communication). Localized structural deformations have been detected biochemically in sharp DNA loops stabilized by Lac repressor (44), lambda repressor (45), and nucleosomes (46), although their mechanical consequences are not known. Many specific kinked structures in protein-DNA complexes are understood at atomic Predictions of the SY theory using these optimal P and C values, extended to longer lengths, compared to our data for short and longer DNAs.…”

Section: Discussionmentioning

confidence: 99%

DNA twisting flexibility and the formation of sharply looped protein–DNA complexes

Cloutier

Widom

2005

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

196

244

View full text Add to dashboard Cite

Gene-regulatory complexes often require that pairs of DNA-bound proteins interact by looping-out short (often Ϸ100-bp) stretches of DNA. The loops can vary in detailed length and sequence and, thus, in total helical twist, which radically alters their geometry. How this variability is accommodated structurally is not known. Here we show that the inherent twistability of 89-to 105-bp DNA circles exceeds theoretical expectation by up to 400-fold. These results can be explained only by greatly enhanced DNA flexibility, not by permanent bends. They invalidate the use of classic theories of flexibility for understanding sharp DNA looping but support predictions of two recent theories. Our findings imply an active role for DNA flexibility in loop formation and suggest that variability in the detailed helical twist of regulatory loops is accommodated naturally by the inherent twistability of the DNA.activation ͉ gene regulation ͉ repression D ouble-stranded DNA in vivo is often sharply distorted away from its classic B-form conformations. In many prokaryotic gene-regulatory complexes, short stretches of DNA, Ϸ100 bp in length, are bent sharply into circular or antiparallel (U-shaped or teardrop-shaped) loops (1, 2). Looping allows synergy between proteins bound at distant DNA sites (3) and decreases the statistical noise in their occupancy (4). DNA looping is also important in eukaryotic regulatory systems (5-7). A striking recent example is the discovery of ligand-dependent DNA looping by the RXR receptor, which may play an important regulatory role at as many as 172 different locations, genome wide, in the mouse (8). In addition, most (Ϸ75-80%) of the length of eukaryotic genomic DNA is sharply bent into nucleosomes (80-bp superhelical loops) (9), which regulate the accessibility and proximity of other DNA-functional sites (10, 11).Prokaryotic and eukaryotic regulatory complexes involving short DNA loops (12-18) place strong constraints on the helical twist of the looped DNA (2). For two DNA-bound proteins to interact when they are separated along the DNA, the proteinbinding sites need to occur on mutually compatible faces of the DNA double helix. This requirement is satisfied by a set of lengths for the intervening DNA that differ from one another by integral multiples of the DNA helical repeat, Ϸ10.5 bp. When the exact length of the intervening DNA in such complexes is suboptimal, the DNA may be under-or overtwisted to allow the protein-protein interaction. The DNA helical twist is altered in other biological systems as well, most notably in the nucleosome, in which the wrapped DNA is under-or overtwisted for most of its length (9).DNA bending and twisting deformations that are required for protein-DNA complex formation come at a cost in free energy, which contributes importantly to the net stabilities and functions of the resulting complexes. For these reasons, the inherent bendability and twistability of DNA itself have been a focus of experimental and theoretical investigation. Classic studies revealed double-st...

show abstract

“…134,129,138 Interestingly, these experiments reveal an increase in the affinity of the Lac repressor to a single site showing that negative supercoiling favors binding. Most importantly though, a dramatic increase in the looping probability was observed.…”

mentioning

confidence: 89%

Biological consequences of tightly bent DNA: The other life of a macromolecular celebrity

et al. 2006

View full text Add to dashboard Cite

The mechanical properties of DNA play a critical role in many biological functions. For example, DNA packing in viruses involves confining the viral genome in a volume (the viral capsid) with dimensions that are comparable to the DNA persistence length. Similarly, eukaryotic DNA is packed in DNA-protein complexes (nucleosomes) in which DNA is tightly bent around protein spools. DNA is also tightly bent by many proteins that regulate transcription, resulting in a variation in gene expression that is amenable to quantitative analysis. In these cases, DNA loops are formed with lengths that are comparable to or smaller than the DNA persistence length. The aim of this review is to describe the physical forces associated with tightly bent DNA in all of these settings and to explore the biological consequences of such bending, as increasingly accessible by single-molecule techniques.

show abstract

DNA supercoiling promotes formation of a bent repression loop in lac DNA

Cited by 239 publications

References 32 publications

Modeling the Lac repressor-operator assembly: The influence of DNA looping on Lac repressor conformation

Modeling the Lac repressor-operator assembly: The influence of DNA looping on Lac repressor conformation

DNA twisting flexibility and the formation of sharply looped protein–DNA complexes

Biological consequences of tightly bent DNA: The other life of a macromolecular celebrity

Contact Info

Product

Resources

About