The three-dimensional solution structure of the 40 residue amyloid beta-peptide, Abeta(1-40), has been determined using NMR spectroscopy at pH 5.1, in aqueous sodium dodecyl sulfate (SDS) micelles. In this environment, which simulates to some extent a water-membrane medium, the peptide is unstructured between residues 1 and 14 which are mainly polar and likely solvated by water. However, the rest of the protein adopts an alpha-helical conformation between residues 15 and 36 with a kink or hinge at 25-27. This largely hydrophobic region is likely solvated by SDS. Based on the derived structures, evidence is provided in support of a possible new location for the transmembrane domain of Abeta within the amyloid precursor protein (APP). Studies between pH 4.2 and 7.9 reveal a pH-dependent helix-coil conformational switch. At the lower pH values, where the carboxylate residues are protonated, the helix is uncharged, intact, and lipid-soluble. As the pH increases above 6. 0, part of the helical region (15-24) becomes less structured, particularly near residues E22 and D23 where deprotonation appears to facilitate unwinding of the helix. This pH-dependent unfolding to a random coil conformation precedes any tendency of this peptide to aggregate to a beta-sheet as the pH increases. The structural biology described herein for Abeta(1-40) suggests that (i) the C-terminal two-thirds of the peptide is an alpha-helix in membrane-like environments, (ii) deprotonation of two acidic amino acids in the helix promotes a helix-coil conformational transition that precedes aggregation, (iii) a mobile hinge exists in the helical region of Abeta(1-40) and this may be relevant to its membrane-inserting properties and conformational rearrangements, and (iv) the location of the transmembrane domain of amyloid precursor proteins may be different from that accepted in the literature. These results may provide new insight to the structural properties of amyloid beta-peptides of relevance to Alzheimer's disease.

Complete sequence-specific, proton-resonance assignments have been determined for the calcium phosphate-stabilizing tryptic peptide beta-casein-(1-25) containing the phosphorylated sequence motif Ser(P)(17)-Ser(P)-Ser(P)-Glu-Glu(21). Spectra of the peptide have been recorded, in separate experiments, in the presence of excess ammonium ions, sodium ions and calcium ions, and of the dephosphorylated peptide in the presence of excess sodium ions. We observed significant changes to chemical shifts for backbone and side-chain resonances that were dependent upon the nature of the cation present. Medium-range nuclear Overhauser effect (nOe) enhancements, characteristic of small structured regions in the peptide, were observed and also found to be cation dependent. The secondary structure of the peptide was characterized by sequential and medium-range (i, i+2/3/4, which denotes an interaction between residue i and residue i+2, i+3 or i+4 in the peptide) nOe connectivities, and Halpha chemical shifts. Four structured regions were identified in the calcium-bound peptide: residues Arg(1) to Glu(4) were involved in a loop-type structure, and residues Val(8) to Glu(11), Ser(P)(17) to Glu(20) and Glu(21) to Thr(24) were implicated in beta-turn conformations. Comparison of the patterns of medium-range nOe connectivities in beta-casein-(1-25) with those in alpha(S1)-casein-(59-79) suggest that the two peptides have distinctly different conformations in the presence of calcium ions, despite having a high degree of sequential and functional similarity.

2D NMR spectroscopic methods have been used to determine the structure of the adduct formed between the antitumor antibiotic hedamycin and the oligodeoxyribonucleotide duplex d(CACGTG)2. Evidence for both intercalation and alkylation in the adduct was observed, and a model for the binding interaction was constructed based on intermolecular NOEs and distance-restrained molecular dynamics. In our computationally refined model, the anthrapyrantrione chromophore of hedamycin is intercalated between the 5'-CG-3' bases with the two aminosugar groups placed in the minor groove and the six carbon bisepoxide side chain located in the major groove. The anglosamine sugar attached at C8 is oriented in the 3' direction relative to the intercalation site, while the N,N-dimethylvancosamine attached at C10 is oriented to the 5' side, with each aminosugar wedged between a guanine exocyclic amino group and one of the groove walls. The terminal epoxide carbon C18 is covalently bound to the N7 atom of the central guanine, as evidenced by lability of the C8 hydrogen of this purine upon reaction with hedamycin. Our binding model places the C10-attached N,N-dimethylvancosamine of hedamycin in van der Waals contact with the alkylated strand. A strong NOE contact verifies the close proximity of the terminal methyl group (C19) of the bisepoxide side chain to the methyl group of the thymine on the 3' side of the alkylated guanine. This, in conjunction with other data, suggests hydrophobic interactions between the bisepoxide chain and the floor of the major groove may contribute to sequence recognition. Furthermore, it is proposed that the 5'-CGT sequence selectivity of hedamycin arises, in part, from complementarity in shape between the chromophore substituents and the major and minor groove at the binding site.