1H NMR, mechanism, and mononuclear oxidative activity of the antibiotic metallopeptide bacitracin: the role of D-Glu-4, interaction with pyrophosphate moiety, DNA binding and cleavage, and bioactivity.

The peptidyl antibiotic bacitracin (Bc) is one of the most widely used antibiotics which can bind divalent transition metal ions, including Mn(II), Co(II), Ni(II), Cu(II), and Zn(II). The metal binding is essential for its antimicrobial activity. Previous analysis of the hyperfine-shifted (1)H NMR signals of Co(II)-Bc A(1) revealed the structure of the metal binding environment and a potential hydrophobic site important for the bioactivity of this antibiotic. Co(II)-Bc in DMSO shows relatively sharper hyperfine-shifted (1)H NMR signals compared with the spectrum acquired in an aqueous solution, allowing more thorough analysis of the signals with 1D and 2D NMR methods. Pyrophosphate and derivatives bind to Co(II)-Bc to form kinetically inert ternary complexes. The coordinated D-Glu-4 is found detached from the metal center of metallobacitracin upon trimetaphosphate binding, implying its role in the antibiotic activity of Bc. We further demonstrate in this report the structure-function relationship on desamido-Bc of low antibiotic activity by the use of NMR, wherein D-Glu-4 is suggested to be important for the bioactivity of Bc. The interaction of the phospho-moiety with Bc is also reflected by DNA binding, wherein metal-free Bc does not bind DNA, whereas various metal complexes of Bc do. Cu(II)-Bc was further demonstrated to bind and oxidatively cleave DNA under reduction conditions in the air. It also exhibited a significant oxidative activity toward catechol oxidation, showing enzyme-like saturation kinetics with k(cat) = 7.0 x 10(-3) s(-1) and k(cat)/K(m) = 2.1 M(-1) s(-1) aerobically and k(cat) = 0.38 s(-1) and k(cat)/K(m) = 14.7 M(-1) s(-1) in the presence of 32 mM of H(2)O(2). The binding of pyrophosphate moiety to metallobacitracin, the detachment of d-Glu-4, and the significant oxidative activity of Cu(II)-Bc provide further insights into the bioactivity of this metallopeptide and Cu-oxygen chemistry.

Synthesis, properties, and NMR studies of a C8-phenylguanine modified oligonucleotide that preferentially adopts the Z DNA conformation.

Carcinogenic aryl hydrazines produce C8-arylated purine adducts. The effect of these adducts on DNA conformation and their role in hydrazine carcinogenesis are unknown. Here, we describe a new synthetic route to produce these adducts that is also compatible with the synthesis of the corresponding phosphoramidites needed for oligonucleotide synthesis. Two oligonucleotides were prepared, an unmodified oligonucleotide, d((5)(')CGCGCGCGCG(3)(')), and a C8-phenylguanine modified oligonucleotide, d((5)(')CGCGCGCGCG(3)(')) (G = 8-phenylguanine). These oligonucleotides were compared using thermal denaturation, circular dichroism, NMR, and molecular modeling. The phenyl modification destabilizes the B DNA form and stabilizes the Z DNA form such that the B:Z ratio is near one under physiological conditions. In light of recent studies that show a role for Z DNA in gene expression and cell transformation, Z DNA stabilization by C8-arylguanine formation from aryl hydrazines may be relevant to their role in carcinogenesis.

Metal binding and structure-activity relationship of the metalloantibiotic peptide bacitracin.

Bacitracin is a widely used metallopeptide antibiotic produced by Bacillus subtilis and Bacillus licheniformis with a potent bactericidal activity directed primarily against Gram-positive organisms. This antibiotic requires a divalent metal ion such as Zn(2+) for its biological activity, and has been reported to bind several other transition metal ions, including Mn(2+), Co(2+), Ni(2+), and Cu(2+). Despite the widespread use of bacitracin since its discovery in the early 1940s, the structure-activity relationship of this drug has not been established and the coordination chemistry of its metal complexes was not fully determined until recently. This antibiotic has been suggested to influence cell functioning through more than one route. Since bacterial resistance against bacitracin is still rare despite several decades of widespread use, this antibiotic can serve as an ideal lead for the design of potent peptidyl antibiotics lacking bacterial resistance. In this review, the results of physical (including NMR, EPR, and EXAFS) and molecular biological studies regarding the synthesis and structure of bacitracin, the coordination chemistry of its metal derivatives, the mechanism of its antibiotic actions, its influence on membrane function, and its structure and function relationship are discussed.

NMR Study of Dendrimer Structures Using Paramagnetic Cobalt(II) as a Probe.

Cobalt(II) has been utilized as an external paramagnetic (1)H NMR probe for the study of the structure of dendrimers that possess specifically located metal recognition sites. The isotropically shifted (1)H NMR signals of the Co(II) complexes of two 2,6-diamidopyridine-containing dendrimers have been fully assigned by means of 1D and 2D NMR techniques, including NOE difference, EXSY, COSY, and TOCSY. T(1) values of the isotropically shifted signals were used to calculate metal-proton distances to build a molecular model of the internal structure of the dendrimers. The presence of sizable cavities within the dendrimers was observed, including a loosely packed conformation for the 2,6-diamidopyridine moiety to bind to potential guest molecules.