Environmentally induced conformational changes in B-type DNA: comparison of the conformation of the oligonucleotide d(TCGCGAATTCGCG) in solution and in its crystalline complex with the restriction nuclease EcoRI

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Abstract
Raman spectroscopic analysis of the secondary structure of the crystalline restriction endonuclease EcoRI, the oligonucleotide d(TCGCGAATTCGCG) in solution, and the corresponding crystalline EcoRi-oligonucleotide complex reveals structural differences between the complexes and uncomplexed protein and oligonucleotide components that appear to be linked to complex formation. Structural differences that are spectroscopically identified include (1) an increase in the population of furanose rings adopting the C3''-endo conformation and (2) spectroscopically observed changes in base stacking which are probably associated with the crystallographically observed distortion of the phosphate backbone about positions C(3)-G(4) and C(9)-G(10) and unwinding between the symmetry-related segments GAA-TTC which make up the central recognition core (McClarin et al., 1986). Changes in base stacking due to distortions and unwinding along the oligonucleotide result in differences in the base vibrational region between the spectra of the complex and the oligonucleotide in solution. The spectroscopic analysis indicates that the C2''-endopopulation is similar for the oligonucleotide in solution and in the complex. The additional C3''-endo population in the complex appears to arise from the conversion of rings adopting alternative conformations such as C1''-exo and O1''-endo. Analysis of the vibrational bands derived from guanine indicates that the population of guanine residues associated with furanose rings in a C2''-endo conformation is similar for the oligonucleotide in solution and in the crystalline complex. This implies that the increase in C3''-endo population is not associated with guanine residues. Large conformational distortions such as those observed in the crystal are not observed for the oligonucleotide in solution. Furthermore, Raman evidence is presented that these distortions are not observed in either the crystal or the solution of the oligomer d(CGCGAATTCGCG). These data suggest that static distortions such as those observed in the crystal are not employed for initial sequence recognition. They appear either as a secondary response to interaction with the protein or as transient fluctuations which exist at a very low level in solution.