Distamycin-induced inhibition of homeodomain-DNA complexes.
The mobility shift assay was used to study the competition of the minor groove binder distamycin A with either an Antennapedia homeodomain (Antp HD) peptide or derivatives of a fushi tarazu homeodomain (ftz HD) peptide for their AT-rich DNA binding site. The results show that distamycin and the homeodomain peptides compete under the conditions: (i) preincubation of DNA with distamycin and subsequent addition of HD peptide; (ii) simultaneous incubation of DNA with distamycin and HD peptide; and (iii) preincubation of DNA with HD peptide and subsequent addition of distamycin. There is also competition when using a peptide which lacks the N-terminal arm of ftz HD that is involved in contacts in the minor groove. It is proposed that the protein's binding affinity is diminished by distamycin-induced conformational changes of the DNA. The feasibility of the propagation of conformational changes upon binding in the minor groove is also shown for the inhibition of restriction endonucleases differing in the AT content of their recognition site and of their flanking DNA sequences. Thus, it is demonstrated that minor groove binders can compete with the binding of proteins in the major groove, providing an experimental indication for the influence of biological activities exerted by DNA ligands binding in the minor groove.
Pharma Division, F. Hoffmann-La Roche Ltd, Basel, Switzerland.
A
A
Dorn
M
M
Affolter
M
M
Müller
W J
WJ
Gehring
W
W
Leupin
D000818:Animals; D051943:Antennapedia Homeodomain Protein; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004214:Distamycins / Q000378:metabolism*; D004330:Drosophila / Q000235:genetics*; D029721:Drosophila Proteins; D051717:Fushi Tarazu Transcription Factors; D005801:Genes, Homeobox / Q000235:genetics*; D018398:Homeodomain Proteins; D007301:Insect Hormones / Q000235:genetics*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D009690:Nucleic Acid Conformation; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D014157:Transcription Factors
D051943:Antennapedia Homeodomain Protein; C497974:Antp protein, Drosophila; D004268:DNA-Binding Proteins; D004214:Distamycins; D029721:Drosophila Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D007301:Insect Hormones; D009687:Nuclear Proteins; D014157:Transcription Factors; C497169:ftz protein, Drosophila; C030000:stallimycin; D015252:Deoxyribonucleases, Type II Site-Specific