JOURNAL OF MOLECULAR RECOGNITION, VOL. zyxwvut 7, 141-156 (1994) zyxwvu REVIEW PAPER zyxwvu Analysing DNA Complexes by Circular and Linear Dichroism Bengt Norden Department of Physical Chemistry, Chalmers University of Technology, S-142 96 Gothenburg, Sweden Tomas Kurucsev Department of Chemistry, University zyxwvutsrq of Adelaide, Adelaide zyxwvutsr 5005, South Australia The application of linear and circular dichroism (LD and CD) in nucleic acid research is illustrated by recent results aimed at answering specific structural problems in the interaction of DNA with molecules of biological importance. We first consider the circumstances under which ligands, such as DAPI (4‘,6-diamidino-2- phenylindole), change their preferred binding mode in the minor groove to major groove binding or intercala- tion. As an extension of this problem we refer to the switch between groove binding and intercalation of structurally similar ligands such as ellipticines and trigonal ruthenium complexes. We also explore the use of LD and CD in the determinationof the structure of the complex formed between the polynucleotidepoly(dA) and the novel ‘peptide nucleic acid’, consisting of nucleic acid bases joined by a polyamide homomorphous with the deoxyribose-phosphatebackbone of DNA. Finally, the structure and interaction of the recombination enzyme RecA with DNA is discussed, in particular the influence of the presence of intercalators, groove binders or covalent DNA adducts. zyxwvutsr Keywords: DNA binding geometries DNA-drug complex DNA-protein complex __ Circular dichroism Linear dichroism Introduction Circular and linear dichroism spectroscopies (CD and LD) may forcefully complement each other and sup- port other branches of spectroscopy, as tools for the study of the structure, interactions and of the function of DNA complexes in solution. Interpretation of dichroic spectra is optimised if the relative magnitudes and directions of the transition dipoles of the chromo/ phores responsible for the light absorption are known. Such information may be obtained experimentally, for instance from stretched film and fluorescence anisotropy measurements, or may be estimated from quantum mechanical calculations. Flow linear dichroism as a spectroscopic tool for characterising DNA conformation (base inclination), flexibility (ability to orient) and binding geometries of DNA-drug and DNA-protein complexes has been recently reviewed in detail (Norden zyxwvutsr et zyxwvut af., 1992a). This is an extension of that review, based on more recent work from our laboratories focusing on three topics of structural characterisation involving the use of LD and CD: (1) Exploration of the criteria that influence and deter- (2) DNA-hybridisdation of a novel group of DNA (3) The binding to DNA of recombinase RecA of mine the binding mode of molecules to DNA. analogues, the peptide nucleic acids (PNA). Escherichia coli. Spectroscopic methods Linear dichroism Linear dichroism, LD, is defined as the differential absorption of light polarised parallel (Apar) and perpen- dicular (Aper) to some laboratory reference axis, at a given wavelength, zyxw 1: LD(I1) =Apar(I1) -A,&). The reduced dichroism (LD‘) is defined as the ratio LD(I1)/A,w(A) where A&) is the (isotropic) absor- bance of the sample. LD can be observed only provided at least partial alignment of the molecules to be studied can be achieved, e.g. through the application of an electric field (electric dichroism) or a hydrodynamic field (flow dichroism). LD is determined relative to a laboratory reference axis taken by convention to be the direction of the orienting field. Our experiments will refer to flow LD which has the advantage over electric LD of being able to use physiological salt concentra- tions and, in addition, of being able to record the whole LD spectrum at a constant, steady-state orientation. Conversely, electric dichroism has the advantage of producing, in general, higher degrees of molecular orientation and providing the possibility of monitoring the process of orientational relaxation when the electric field is switched off. The quantity L D is related to the degree of orient- ation, described by an orientation factor S (S= 1 for perfect alignment of the DNA helix parallel to the flow, S=O for random orientation), and to the angle, a, between the helix and the light-absorbing transition dipole. Provided there is no overlap between the absorption bands of the oriented molecule one has (Norden et al., 1992a): LDr=LD(I1)/A,s,(A) =S X 3/2 x (3 COS’ a - 1) (1) The most common applications of LD to structural studies involving DNA are illustrated in a simplified way in Fig. 1. The LD spectrum of B-DNA shown in the central diagram (marked B) is negative at the lowest wavelength region and of the same shape as the Received IS August I993 CCC 0952-3499/94/020141- 16 zyxwvutsr 0 1994 by John Wiley & Sons, Ltd.