Carbohydrate-Minor Groove Interactions in the Binding of Calicheamicin .gamma.1I to Duplex DNA

We report studies of the contribution of DNA structure, holding the sequence constant, to the affinity of calicheamicin ␥ 1 I and its aryltetrasaccharide moiety for DNA. We used polynucleotide chains as models of known proteinbinding sequences [the catabolite activator protein (CAP) consensus sequence, AP-1 and cAMP response element (CRE) sites] in their free and protein-bound forms. The proteins were selected to provide examples in which the minor-groove binding site for the carbohydrate is (CAP) or is not (GCN4) covered by the protein. Additionally, peptides related to the GCN4 and CREB families, which have different bending effects on their DNA-binding sites, were used. We observe that proteins of the CREB class, which induce a tendency to bend toward the minor groove at the center of the site, inhibit drug-cleavage sites located at the center of the free AP-1 or CRE DNA sites. In the case of GCN4, which does not induce DNA bending, there is no effect on calicheamicin cleavage of the CRE site, but we observe a GCN4-induced rearrangement of the cutting pattern in the AP-1 site. This effect may arise from either a subtle local conformational rearrangement not accompanied by bending or a localized reduction in DNA f lexibility. Whereas GCN4 binding is not inhibited by the calicheamicin aryltetrasaccharide, binding of CAP to its DNA target is significantly inhibited, and calicheamicin cutting of DNA at the center of the CAP-DNA complex site is strongly reduced by protein binding. This result probably ref lects steric inhibition of drug binding by the protein.

Section: Discussionmentioning

confidence: 70%

Interaction of calicheamicin γ ₁ ^I and its related carbohydrates with DNA–protein complexes

Sissi

Aiyar²,

Boyer³

et al. 1999

“…Further, the amino group of the 5'-guanine in the d(A-G-G-A) segment is a critical recognition element since its replacement by inosine (lacking a 2-amino group) results in a greatly reduced binding to calicheamicin y1I (18). These constraints are met in our solution structures of the complexes that show direct DNAto-ligand hydrogen bonding interaction between the exposed amino proton of G18 and the iodine atom of calicheamicin.…”

Section: (T-c-c-t)d(a-g-g-a) Minor Groove Binding Site and Exhibits mentioning

confidence: 88%

“…Schreiber and colleagues (17) (18). Further, the amino group of the 5'-guanine in the d(A-G-G-A) segment is a critical recognition element since its replacement by inosine (lacking a 2-amino group) results in a greatly reduced binding to calicheamicin y1I (18).…”

Section: (T-c-c-t)d(a-g-g-a) Minor Groove Binding Site and Exhibits mentioning

confidence: 99%

Calicheamicin-DNA complexes: warhead alignment and saccharide recognition of the minor groove.

Ikemoto

Kumar

Tao

et al. 1995

“…The (15). This fact is also reflected in the results of the in vitro transcription test where a template with the TCCT sites mutated to TGGT shows comparable inhibition at a 2-to 4-fold higher carbohydrate concentration.…”

Section: Discussionmentioning

confidence: 86%

“…The carbohydrate portion of calicheamicin has been obtained by total synthesis, and its interaction with DNA has been studied extensively (12)(13)(14). One of the preferred target sequences is the tetranucleotide TCCT (3); others include TCTC and TTTT (2,10,15). Structural and chemical analysis has shown that the carbohydrate tail of calicheamicin is oriented toward the 3' end of the TCCT surface and that the iodine of the aryltetrasaccharide plays a critical role in the sequence-selective recognition of DNA (2,5,10).…”

mentioning

confidence: 99%

Sequence-selective carbohydrate-DNA interaction: dimeric and monomeric forms of the calicheamicin oligosaccharide interfere with transcription factor function.

Liu

Smith

Ajito

et al. 1996

The synthetic oligosaccharide moiety of the antibiotic calicheamicin and the head-to-head dimer of this oligosaccharide are known to bind to the minor groove of DNA in a sequence-selective manner preferring distinct target sequences. We tested these carbohydrates for their ability to interfere with transcription factor function. The The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.carbohydrates can interfere with the sequence-specific binding of transcription factors to DNA and with the activation of transcription. We find that the dimer of the calicheamicin oligosaccharide is a much more effective inhibitor of biological activity than is the monomer. MATERIALS AND METHODSCalicheamicin Carbohydrates. The aryltetrasaccharide of calicheamicin was obtained by total synthesis as described (12,17). The synthesis of the head-to-head dimer of this compound has also been reported (18). Structure and purity of the carbohydrates were verified by thin-layer chromatography and IH NMR spectroscopy.Electrophoretic Mobility Shift Assays. The oligonucleotides used in electrophoretic mobility shift assays are shown in Table 1. The AP-1 binding site is derived from the human collagenase gene promoter. It does not contain a TCCT sequence and serves as a control for nonspecific binding of the carbohydrate. In the AP-1(TCCT) and the AP-1(TCCT)2 oligonucleotides, TCCT sequences (doubly underlined in Table 1) were constructed near the AP-1 consensus. The AP-1(TCCT) oligonucleotide contains one TCCT site, five nucleotides 5' to the consensus, and the AP-1(TCCT)2 oligonucleotide contains two TCCT sites, one immediately 5' to the consensus, and a second one six nucleotides upstream. AP-1(DM) contains the optimal dimer DNA binding sequence AGGAAGTCCT im-