Screening of threading bis-intercalators binding to duplex DNA by electrospray ionization tandem mass spectrometry.
The DNA binding of novel threading bis-intercalators V1, trans-D1, and cis-C1, which contain two naphthalene diimide (NDI) intercalation units connected by a scaffold, was evaluated using electrospray ionization mass spectrometry (ESI-MS) and DNAse footprinting techniques. ESI-MS experiments confirmed that V1, the ligand containing the -Gly3-Lys- peptide scaffold, binds to a DNA duplex containing the 5'-GGTACC-3' specific binding site identified in previous NMR-based studies. The ligand formed complexes with a ligand/DNA binding stoichiometry of 1:1, even when there was excess ligand in solution. Trans-D1 and cis-C1 are new ligands containing a rigid spiro-tricyclic scaffold in the trans- and cis- orientations, respectively. Preliminary DNAse footprinting experiments identified possible specific binding sites of 5'-CAGTGA-5' for trans-D1 and 5'-GGTACC-3' for cis-C1. ESI-MS experiments revealed that both ligands bound to DNA duplexes containing the respective specific binding sequences, with cis-C1 exhibiting the most extensive binding based on a higher fraction of bound DNA value. Cis-C1 formed complexes with a dominant 1:1 binding stoichiometry, whereas trans-D1 was able to form 2:1 complexes at ligand/DNA molar ratios >or=1 which is suggestive of nonspecific binding. Collisional activated dissociation (CAD) experiments indicate that DNA complexes containing V1, trans-D1, and cis-C1 have a unique fragmentation pathway, which was also observed for complexes containing the commercially available bis-intercalator echinomycin, as a result of similar binding interactions, marked by intercalation in addition to hydrogen bonding by the scaffold with the DNA major or minor groove.
Mazzitelli
Carolyn L
CL
Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas 78712-0165, USA.
Chu
Yongjun
Y
Reczek
Joseph J
JJ
Iverson
Brent L
BL
Brodbelt
Jennifer S
JS
D001665:Binding Sites / Q000235:genetics; D004247:DNA / Q000737:chemistry*; D018983:DNA Footprinting; D007094:Imides / Q000737:chemistry; D007364:Intercalating Agents / Q000737:chemistry*; D008024:Ligands; D008403:Mass Screening; D008970:Molecular Weight; D009281:Naphthalenes / Q000737:chemistry; D009841:Oligonucleotides / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry
D007094:Imides; D007364:Intercalating Agents; D008024:Ligands; D009281:Naphthalenes; D009841:Oligonucleotides; D010455:Peptides; D004247:DNA