Deglyco-bleomycin-iron complexes: implications for iron-binding site and role of the sugar portion in bleomycin antibiotics.

Sir: The oxidation of bleomycin (BLM)-Fe-DNA complex by molecular oxygen leads to damage of DNA.'," This activity is related to the bifunctionality of BLM, namely DNAand iron-binding. The bithiazole and terminal amine residues contribute toward the binding to DNA3,4) and the pyrimidoblamyl-(-hydroxyhistidyl portion (the amino acid residue containing pyrimidine is called pyrimidoblamic acid) is involved in chelation with metal ions.2,5,11 From 1H NMR analysis of a diamagnetic BLM-Fe(II)-CO adduct, the carbamoyl group on the 3-position of mannose was proposed to be one coordination donor group toward the ferrous ion, in conjunction with carbon monoxide, the primary and secondary amine nitrogens, the pyrimidine N-1 and the imidazole N-1.7)However, this model has several drawbacks. 'The proposed 5-5-9 chelate ring member and the coordination of the carbamoyl amide nitrogen are considered to be unlikely, because certain spectroscopic properties8) and the binding of oxygen antagonists (CO, NO, and C2H5NC)9) similar to hemoproteins strongly suggest a rigid chelate conformation of the BLMiron complex. Our potentiometric study showed one proton dissociation (pKc=5.5) from the peptide backbone group in the BLM-Fe(II)-complexation.10) In addition, the ESR results of the BLM-Co(II) complex and its dioxygen adduct revealed that the primary amino nitrogen is the fifth axial donor atom.11) In this paper, deglyco-BLM was used to clarify the role of the sugar portion for the coordination and Oz-activation of "active BLM-Fe(II) complex species". Deglyco-BLM lacks the sugar moiety in BLM Molecule. Deglyco-BLM was obtained from the culture broth of BLM fermentation and also from the mild acid hydrolysate of BL M.12) Recently, the total chemical synthesis of deglyco-BLM-A2 was achieved by us."' The 1 : 1 deglyco-BLMFe(II) complex was prepared by mixing deglycoBLM and Fe(ll) ion in aqueous solution (pH 6.9) containing a small amount of NaBH4 under the condition of total deaeration. The Fe(II)-NO (or CO) adducts of deglyco-BLM and BLM were prepared by addition of a few milligrams of NaNO2 (or CO gas) and sodium borohydride to the Fe(II) complexes. Spin-trapping experiments using N-tert-butyl-a-phenylnitrone were carried out according to the previously reported procedure." X-Band ESR(sample conc.: 1 mM) and 220 MHz FT-1H NMR (10 mM) spectra were