Orientation of Iron Bleomycin and Porphyrin Complexes on DNA Fibers

Abstract

Bleomycin (Blm) is an antitumor agent that requires iron and oxygen for strand cleavage of DNA. In this study, ferric bleomycin, Fe(III)Blm, or the nitric oxide adduct of ferrous bleomycin, ON-Fe(II)Blm, were bound to one-dimensionally oriented DNA fibers. Reductive nitrosylation of Fe(III) complexes took place in situ on B-form DNA fibers. Electron paramagnetic resonance (EPR) spectra were obtained as a function of the angle phi between the magnetic field B and the fiber axis Zf. For comparison, EPR spectra were acquired for ON-Fe(II)TMpyP and ON-Fe(II)TMpyP-Im on oriented DNA fibers, where TMpyP is 5,10,15,20-tetrakis(1-methyl-4-pyridino)porphyrin and Im is imidazole. EPR spectra showed both low-spin Fe(III)Blm and ON-Fe(II)Blm bound to B-form DNA in two slightly different binding orientations in the ratio of 1:0.2. With A-form DNA, a fraction of bound Fe(III)Blm was high spin. Specifically, the angle beta between the fiber axis Zf and the g axis, gz, perpendicular to or nearly perpendicular to the equatorial plane of the iron complex was estimated as 20 degrees and 25 degrees for ON-Fe(II)Blm and 30 degrees and 25 degrees for Fe(III)Blm, respectively. The angle gamma that determines the orientation of gx and gy axes was estimated as 90 degrees for the two ON-Fe(II)Blm species and 10 degrees for the two Fe(III)Blm species, respectively. The NO was held rigidly in place as the temperature increased from 123 K to room temperature for ON-Fe(II)Blm but not for ON-Fe(II)TMpyP or ON-Fe(II)TMpyP-Im. It is hypothesized that the NO is structurally oriented by hydrogen bonding like the peroxide is held in HO2(-)-Co(III)Blm (Wu et al. J. Am. Chem. Soc. 1996, 118, 1281-1294). The EPR parameters are consistent with a six-coordinate complex for ON-Fe(II)Blm, although the superhyperfine structure from the trans nitrogen was not detected. The increase in g value anisotropy upon binding ON-Fe(II)Blm to DNA fiber may be caused by an increase in the overlap of d pi and 2p pi* orbitals induced by an interaction of NO with DNA and/or by a perturbation of d orbitals due to the pyrimidine-guanine interaction. It is concluded that the EPR parameters of ON-Fe(II)Blm and Fe(III)Blm bound to oriented DNA support the hypothesis that FeBlm species bind to DNA with adduct structures similar to those formed by related CoBlm species and DNA.

Deglyco-peplomycin metal complexes on DNA fibers: a role of the sugar moiety for the stability and the orientation of the complexes

“…Figure 4 shows the EPR spectra of Fe(III)BLM and Fe(III)dPEP on A-form DNA fibers at -150°C. In the case of Fe(III)BLM, the intensity of the signal at g=4.27 was considerably increased on fabrication of A-form DNA fibers from the pellet as previously reported [20], suggesting that the change in the conformation of the DNA induced a deformation of the metal-binding moiety to form the high-spin species of Fe(III)BLM. In contrast, the low-spin species of Fe(III)dPEP completely disappeared on the A-form DNA fibers and, instead, a broad F-dependent signal ranging over 400 mT emerged, overlapping with the F-independent signals at g=8.21, 5.31, and 4.27.…”

“…The molar ratio of metal complex to DNA base pairs in the solution was adjusted to 1:25. The solution was stirred overnight at 4°C and then ultracentrifuged at 320,000g for 7 h. The obtained DNA pellet was used for the preparation of DNA fibers as described previously [20,21,22].…”

“…In the case of low-spin Fe(III)BLM, it was easily nitrosylated to ON-Fe(II)BLM [20]. The formation of ON-Fe(II)BLM has been attributed to the presence of a metal-binding moiety in BLM similar to that of porphyrins [4,24].…”

“…We have shown that the orientation of the paramagnetic center of metallo-bleomycin bound to DNA fibers can be assessed by EPR spectroscopy [16,17]. In this article, we report how the sugar group affects the stability and the orientation of the complex on DNA, comparing the EPR spectra of Fe(III), O 2 Co(II), and Cu(II) complexes of BLM and deglyco-peplomycin (dPEP) on DNA fibers.…”

EPR study of the dynamic behavior of cis,cis-1,3,5-triaminocyclohexane Cu(II) complex on B-form DNA-fibers

“…To investigate the binding structure of the complex on DNA, and to get information for the hydrolysis mechanism of phosphate dieseter bond, we have now measured the electron paramagnetic resonance (EPR) spectra of [Cu(TACH)] 2þ on DNA-fibers under different conditions and found a novel dynamic behavior of the complex on the B-form DNA-fibers. The DNA-fiber EPR spectroscopy affords unique information on the stereospecificity and the dynamic properties of paramagnetic complexes on DNA [12][13][14]. We have shown that the efficient hydrolytic reaction catalyzed by [Cu(TACH)] 2þ originates from such novel dynamic behavior of the complex on DNA.…”