Allostery: DNA Does It, TooJonathan B. Chaires*James Graham Brown Cancer Center, University of Louisville, 529 South Jackson Street, Louisville, Kentucky 40202

A llostery is a central concept in bio-chemistry and molecular biology. In-deed, Jacque Monod declared al-

lostery to be “the second secret of life” (1).“Allostery” is derived from the Greek allos(“other”) and stereos (“space” or “shape”)and is a manifestation of the thermody-namic coupling of binding reactions to con-formational transitions in macromolecules.Allosteric interactions are ubiquitous in theregulation and modulation of enzymaticactivity. The concept of allostery is com-monly presented in introductory biochemis-try textbooks and is typically presentedwith reference to the well-known Monod�

Wyman�Changeux (MWC) and Koshland�

Nemethy�Filmer models. These modelsfeature the triggering of a “switch” betweenconformations of protein subunits within amultisubunit protein complex brought on byan initial binding event, a switch that altersthe affinities for subsequent binding events.The view of allostery has evolved to recog-nize that the fundamental capacity to un-dergo conformational changes in responseto binding may be intrinsic to all proteinsand that such coupled conformational shiftsmay be a fundamental mechanism in regu-lation and communication (2, 3). Allosteryprovides a means of responding to a chang-ing environment to modulate activity. Thepaper by Moretti and co-workers on page220 of this issue shows that allostery is notunique to proteins and that DNA is allo-steric, too (4).

It is not commonly appreciated that DNAis allosteric. DNA is often mistakenly viewedas an inert lattice onto which proteins as-semble to replicate or transcribe genes. Pro-

tein binding might be guided to specificsites on the inert lattice by sequence-specific interactions utilizing recognition el-ements residing in the major or minorgrooves, in particular specific patterns of hy-drogen bond donors and acceptors alongthe edges of base pairs (5). Several excitingrecent developments, including the reportby Moretti, contradict this view of the staticlattice and suggest that DNA may be a moreactive partner in its own replication and tran-scription through allosteric transitions in itsstructure that modulate protein binding.There is also a longer history of allostericDNA that bears remembering.

As early as 1972, the intercalator ethid-ium was shown to act as an allosteric effec-tor and could convert left-handed Z DNA to aright-handed form (6). The MWC model forallostery was invoked to explain highly co-operative binding isotherms observed forethidium binding to Z DNA. A few years later,Crothers and co-workers showed that bind-ing of the groove-binder distamycin inducesa cooperative transition of calf thymus DNAto a new form with higher affinity for thedrug and altered structural properties (7).Subsequently, a statistical mechanicaltheory was presented by the Crothers labo-ratory for the calculation of binding iso-therms when binding is coupled to a DNAstructural change (8). The theory was analo-gous to the MWC model for allosteric bind-ing to proteins but differed in the detailsnecessary to describe the conformationaltransition of the DNA lattice, as illustrated(Figure 1, panel a). The model appliesequally well for protein or small-moleculebinding to DNA. The Crothers model was

*Corresponding author,j.chaires@louisville.edu.

Published online April 18, 2008

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© 2008 American Chemical Society

ABSTRACT Allostery is a central concept for un-derstanding protein function and regulation. It isless well appreciated that DNA is allosteric, too,and that DNA conformational changes can bycoupled to protein binding interactions on theDNA lattice. Allosteric DNA interactions are emerg-ing as important features in the assembly of themolecular machines that regulate transcription.

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subsequently used to quantitatively ana-lyze binding isotherms for the interaction ofthe anticancer drug daunorubicin and its en-antiomer with both left- and right-handedDNA (9, 10). These studies showed thatbinding affinity was enhanced by nearly 40-fold by coupling to the DNA conformationalchange, adding �2 kcal mol�1 more favor-able binding free energy.

A contrasting model for cooperative bind-ing of proteins to a DNA lattice is that ofMcGhee and von Hippel (11), which de-scribes neighbor exclusion effects. In thatmodel, the conformation of the DNA latticeis unchanging, but positive cooperativitycan arise from interactions between boundproteins. Thus, protein binding is tighter atcontiguous sites because of the additionalinteractions between bound proteins(Figure 1, panel b). Binding to a contiguoussite may contribute up to 3�4 kcal mol�1 ofmore favorable binding free energy (12).

The Crothers allosteric and the McGhee�

von Hippel models predict binding iso-therms with distinctly different shapes thatcan be distinguished by nonlinear curve fit-ting of binding isotherms to the respectivemodels (13).

Another key historical concept is that of“telestability” (14, 15). Pioneering studiesfrom the Wells laboratory showed that thesequence of one region of a syntheticdouble-helical DNA affected the physicalproperties of a contiguous but remote re-gion. A manifestation of this phenomenonis that the binding affinity to a specific se-quence can be altered by changes in thesurrounding sequences, leading to se-quence context effects. For example, thebinding free energy of the drug actinomycinD for the sequence 5=-AGCT can vary bynearly 1 kcal mol�1, depending on theflanking sequence surrounding the actualbinding site (16).

The transmission of allosteric effects inDNA has been amply demonstrated by ex-periment, and the many examples havebeen reviewed (17, 18). Without question,the binding of proteins or small moleculesto a DNA lattice can produce coupled confor-mational changes that alter the binding ofsubsequent molecules, often over longdistances.

More recent studies have indicated theimportance of DNA allosteric effects in theassembly of the macromolecular machinesused for gene expression and replication.The “enhanceosome”, for example, is a pro-tein complex that binds to the enhancerregion of a gene, either upstream or down-stream of the promoter (19). The enhanceo-some accelerates transcription of the gene.The binding and assembly of the activatingproteins, some of which may be transcrip-tion factors, are cooperative, in part becauseof energetically favorable protein�proteininteractions formed in the complex. Arecent detailed structural analysis of theinterferon-� enhanceosome revealed, how-ever, that cooperative protein�protein inter-actions alone may not be enough to ac-count for the high-precision assembly ofthe machinery (20). A detailed examinationof an atomic model of the interferon-� en-hanceosome led to the conclusion that the“paucity of local protein�protein contactssuggests that cooperative occupancy of theenhancer comes from both binding-inducedchanges in DNA conformation and interac-tions with additional components” (20).This study highlights a significant problem,namely, that it has been difficult, if not im-possible, to separate and quantify the con-tribution of DNA conformational changes tothe assembly process, such as constructionof the enhanceosome. It is this problemthat Moretti and co-workers have attackedand to which they have provided a noveland elegant solution.

Moretti and co-workers explore the coop-erativity in the DNA binding interactions ofHox and Exd proteins. Exd is a transcription

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a

b

Figure 1. Types of cooperativity in protein�DNA interactions. a) Allosteric DNA. DNA can equili-brate between two conformations (yellow and green). Proteins bind to each with different affin-ity. Binding of the first protein shifts the DNA conformational equilibrium to a higher affinityform, facilitating the binding of subsequent proteins. Additional favorable binding energy couldarise from protein�protein interactions between molecules bound to the lattice. b) Nonallostericbinding. The conformation of the DNA lattice is constant. Binding of a protein to a contiguoussite next to another bound protein leads to favorable protein�protein interactions, facilitatingbinding of the second molecule.

208 VOL.3 NO.4 • 207–209 • 2008 www.acschemicalbiology.orgCHAIRES

factor that guides Hox to unique DNA bind-ing sites. Exd binds to DNA with nearly 4kcal mol�1 more favorable binding free en-ergy in the presence of Ubx (a member ofthe Hox family). A mutant Ubx in which pos-sible protein�protein interactions with Exdare largely eliminated still facilitates bindingby �1 kcal mol�1. Careful scrutiny of theknown crystal structures of the complex sug-gests that a Ubx-induced DNA conforma-tional change facilitates Exd binding, specif-ically a widening and decrease in the depthof the major groove. A hairpin polyamidemolecule was designed that could bind se-lectively into the minor groove with transmit-ted effects into the major groove that mimicthe changes that were seen to be inducedby protein binding. The designed “wedge”molecule was found to enhance Exd bind-ing by 1.5 kcal mol�1 in the absence of Ubx.The wedge thus acts as an allosteric effec-tor to facilitate binding of the protein. The re-sults and strategy are significant becausethey provide a new tool for dissecting theenergetic contributions of coupled DNA con-formational changes to specific protein�

DNA binding interactions. Above all, theresults conclusively show that DNA is allo-steric, too, and that the effects of suchallostery cannot be ignored.

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