Nucleic Acids Research, 1992, Vol. 20, No. 4 653-657
© 1992
CHEMISTRY |
Synthesis of isotope labeled oligonucleotides and their use in an NMR study of a protein-DNA complex
Department of Chemistry, University of Utrecht Padualaan 8, 3584 CH Utrecht 1Gorlaeus Laboratory, University of Leiden PO Box 9502, 2300 RA Leiden, The Netherlands
*To whom correspondence should be addressed
Received January 3, 1992. Revised January 28, 1992. Accepted January 28, 1992.
The synthesis of an oligonucleotlde labeled with 13C at the thymine methyls and 15N at the exocycllc amino groups of the cytoslnes Is described. 13CH3I and 15NH4OH were used as sources of the labels. The labeled ollgonucleotide was characterized by several NMR techniques. The duplex possesses a labeled functional group in the major groove at every base pair which makes it a very suitable probe for the study of sequence-specific protein-DNA interaction. The labeled thymine methyl group facilitates the detection of hydrophobic contacts with aliphatic side-chains of proteins. This is demonstrated in an NMR study of a complex between the glucocorticold receptor DNA-bindlng domain and the labeled ollgomer, which revealed a hydrophobic contact between a thymine methyl group and the methyl groups of a valine residue. There are indications for small differences between the solution structure the X-ray structure of the complex.