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Published August 3, 1999 | public
Journal Article

Perturbing the DNA Sequence Selectivity of Metallointercalator−Peptide Conjugates by Single Amino Acid Modification

Abstract

Metallointercalator−peptide conjugates that provide small molecular mimics to explore peptide−nucleic acid recognition have been prepared. Specifically, a family of peptide conjugates of [Rh(phi)_2(phen')]^(3+) [where phi = 9,10-phenanthrenequinone diimine and phen' = 5-(amidoglutaryl)-1,10-phenanthroline] has been synthesized and their DNA-binding characteristics examined. Single amino acid modifications were made from the parent metallointercalator−peptide conjugate [Rh(phi)_2(phen')]^(3+)-AANVAIAAWERAA-CONH_2, which targets 5'-CCA-3' site-specifically. Moving the glutamate at position 10 in the sequence of the appended peptide to position 6 {[Rh(phi)_2(phen')]^(3+)-AANVAEAAWARAA-CONH_2} changed the sequence preference of the metallointercalator−peptide conjugate to 5'-ACA-3'. Subsequent mutation of the glutamate at position 6 to arginine {[Rh(phi)_2(phen')]^(3+)-AANVARAAWARAA-CONH_2} caused more complex changes in DNA recognition. Thermodynamic dissociation constants were determined for these metallointercalator−peptide conjugates by photoactivated DNA cleavage assays with the rhodium intercalators. At 55 °C in the presence of 5 mM MnCl_2, [Rh(phi)_2(phen')]^(3+)-AANVAIAAWERAA-CONH_2 binds to a 5'-CCA-3' site with K_d = 5.7 × 10^(-8) M, whereas [Rh(phi)_2(phen')]^(3+)-AANVAEAAWARAA-CONH_2 binds to its target 5'-ACA-3' site with K_d = 9.9 × 10^(-8) M. The dissociation constant for [Rh(phi)_2(phen')]^(3+) with random-sequence DNA is 7.0 × 10^(-7) M. Structural models have been developed and refined to account for the observed sequence specificities. As with much larger DNA-binding proteins, with these metal−peptide conjugate mimics, single amino acid changes can lead to single or multiple base changes in the DNA site targeted.

Additional Information

© 1999 American Chemical Society. Received August 24, 1998; Revised Manuscript Received June 4, 1999. Publication Date (Web): July 16, 1999. We are grateful to the NSF (Grant CHE-9422547) for financial support of this work. C.A.H. thanks W. R. Grace & Co. for a summer research fellowship. We thank the Biopolymer Synthesis and Analysis Center at Caltech for their technical assistance.

Additional details

Created:
August 19, 2023
Modified:
October 18, 2023